BP Summaries

Microscopy Summary
Light (.2 m)
Condenser: focuses light
Objective: resolution
Ocular: magnifies
No screen (only in EM)
SEM (2 nm)
Light source = elec. beam
Surface only
No obj lens (doesnt penetrate)
TEM (.2 nm)
Penetrates
Expensive
Methods (TEM & light)
Fixation (glutraldehyde), dehydration
(EtOH), Infiltration/embedding (Epon
812 resin), section (diamond knife),
staining (heavy metals for TEM)
Freeze-fracture: fixation (optional),
freezing, fracturing/etch,
coating/replication (mold) avoids
artifacts of chemical
Common dyes: Hematoxylin (basic dye-
stains acids blue); eosin (stains bases
red); PAS (carbs); Sudan Black (lipids)
Common metals: uranyl acetate, the
lead citrate for TEM
Markers
Antibodies
polyclonal: many bands, quick;
monoclonal: need 1 specific
antibody
Direct method: 1 antibody, short
life, $$$. Indirect method: 2
antibody marked.
Flourecence
FITC: green . Rhodamine: red
Heavy metal
Gold colloid
Enzyme
Catalse: DAB method- turns black
with osmium


DNA Summary
Prokary. Genome
Circular, DS, no membrane
Mitochondria similar, but import hosts
enzymes
Viruses
Retroviruses: use rev transcriptase to
make DS DNA from RNA, insert into host.
Orthomyxovirus: make only RNA , not
DNA
DNA Structure
Nucleotides: base, sugar, phosphate. (
Sides: no P)
Chargaffs Rule: A-T (2 H-bonds) G-C (3
bonds)
DS helix: antiparallel; B-form common
(major/minor groves); phosphodiester
bonds.
Chromosome structure
Nucleosome: DNA around histone (H1,
H2ab, H3, H4 subunits)
Heterochromatin: tightly wound
Euchromatin: exposed, cleaved by
DNAse.
P arm small. Q arm long. Centromere
center.




DNA replication (Eukary)
Semiconservative: 1 strand original,
1 strand built anew
Enzymes:
Gyrase/topioisomerase: stop
supercoiling
DNAses: cleave phosphod bonds
Helicase: unzip DNA (form
replication fork).
Lagging strand: Okazaki
Primase: put down RNA primer
DNA pol: read in 3-5 direction
(strand built in 5-3) . Use dNTP
as NRG.
Pol III: Prokary. Has only 3 to 5
exonuclease activity during repair.
Pol : Eukary.
Exonuclease (part of DNA Pol):
5 -remove primer. 3 -remove
single-base errors
Ligase: close gaps btw Okazaki
fragments
Final steps
Eukary genomes arent circular,
have telomeres. Telomerase
extends 5 ends of original strand
(rev. transcribes RNA primer) .
Long telomere folds over to cap
3 end of new short strand
Review anti-caner drugs!
Acid/Base Summary
Acid: proton donor
pH = -log[H
+
]
HH equation: pH = pK + log [A
-
/HA]
Buffer: mixture of weak acid and its salt. Adding some acid pushes rxn to make more
salt.
Buffering zone: pH changes very little. Narrow! (+/- 1 pk unit)
Best buffering: pK = pKa
Physiological Buffers:
Bicarbonate: metabolic component.
CO
2:
Respiratory component
Eg: Resp. acidosis: increase in CO
2
, body compensates by inc. bicarb.
Note: Alkalosis is rare.



Carb/Lipid Summary
Chirality: image cant be superimposed
Enantiomers: mirror images
Diasteromers: not mirror images
Carbs
D-isomer: -OH is right of highest number
chiral carbon
5 & 6 chain carbs can form cyclic
hemiacetals in solution
Hawethorne: : -OH is down
Glucose ring: pyranose. Fructose ring:
furanose
See notes for illustration
Straight chains are reducing sugars
Glycosidic linkage: forms disaccharides


Lipids
CH
3
(CH
2
)
n
COOH
Carbon 1: COOH
Carbon 2 (): (CH
2
)COOH
Unsaturated: at least 1 cis double
bond (Hs on same side)
CH
3
(CH
2
)
4
CH = CH CH
2
CH =
CH(CH
2
)
7
COOH

9, 12
octadecadienoic acid
Polyunsaturated: numbered
from the CH
3
end.
Triglyceride: glycerol esterified
with 3 fatty acid chains
Others: Phospholipids (cell
membrane), steroids


Amino Acid Summary
AA properties:
Most are L (-NH
3
to left)
Zwitterion: + and charge. Makes aa
more soluble, compatible w/ life
(melting pts, etc)
Isoelectric pt: where aa has no
charge
Peptide bonds form chains (N-
terminus to C-terminus). Stable &
planar.
1 structure: aa sequence
2: sheets, helices, random coils
3: how prot folds (hydrophobic,
electrostatic, disulfide interactions)
4: subunits
Some names:
Asparginine-N; Aspartic acid-D;
Glutamine-Q; Lysine-K
Cleavage (Enzyme/chemical)
Trypsin: cleaves C-end of Arg and
Lys
GLY ARG SER -- GLY ARG + SER
Chymotrypsin: cleaves aromatics
Phe, Tyr, Try
Pepsin: same as above, also Met
and Leu
CNBr: cleaves Met

Protein Characterization
Gel electro: separates proteins by
charge (neg. ones move towards
pos. anode)
SDS-page: molecular weight
Isoelectric: pH (stops migrating when
it hits pI)
Ion-exchange chromatography
CM-cellulose: neg. ligand,
pushes - prot out column, +
remain (cation exchanger)
DEAE-cellulose: anion exchanger
Gel-filtration (smaller prot caught in
gel, bigger ones elute out 1
st
)
Affinity chromatography: antibodies
are stationary phase ($$$!)
Spectroscopy methods
(fluorescence, mass, etc)
Cell Membrane Summary
Cell membrane structure
Fluid mosaic: integral prot
(amphopathic); peripheral
(hydrophillic); glycoproteins (cell-
cell recognition, neg. charge)
Membrane lipids: phosholipids,
cholesterol. Turn black from
osmium dye
Has tri-laminar appearance
Membrane proteins: transporters,
enzymes, receptors, linkers (hold
cytoskeleton)
Cell membrane fxns
Selectively permeable
Passive transport (down conc.
grad): simple diffusion = small
mlx. Facilitated transport
(through protein channels) =
large or polar
Active transport: Up conc.
Gradient, use NRG (Na/K
ATPase: Na pumped out)
Signal transduction
Transmitter-gated ion channel (ligand
bind, lets ions through); enzyme-lined
receptor (binding opens up active site); G-
protein-linked receptor (amplies signals
w/ GTP)
Endocytosis
Phagocytosis (large things)
Pinocytosis (small dissolved solutes)
Receptor-mediated = receptor proteins w/
target on inside of vesicle, clathrin coat
outside.
Exocytosis
From Golgi
Secretion can be Mediated (by
hormone/neurotransmitter) or Constitutive
(unregulated)
Membrane Flow
Btw. Membrane-bound organelles
After de-coating, V-snare binds to T-snare
on target destination.
Vesicle needs to be 1.5 nm away to fuse.


Cell Organelles
Rough ER
Parallel cisternae, covered w/
ribosomes (hematoxylin dye turns
acid [RNA in ribosomes] dark)
Makes proteins for secretion,
insertion in membrane, fusion to
lysosome
Free ribosome reads signal on
mRNA, signal recognition particle
stops translation until ribosome
docked to ER (by receptor
ribophorin), continues trans.
protein into ER lumen. N-terminus
translated 1
st
. Ribosome detaches
from mRNA strand at completion.
Ribosome also glycosylates
proteins AND inserts
transmembrane proteins w/ help of
translocon.
Plasma cells, liver, pancreas,
fibroblasts, osteo/chrondocytes &
prot storing cells rich in RER
Smooth ER
Tubular cisternae, no ribosomes. Not
dyed dark.
Makes steroids and glycogen,
metabolises lipids, stores AND
relaeases Ca ions, drug detox
Steroid-secreting cells (adrenals,
gonads), active metabolic cells
(liver/pancreas) and muscle rich in SER
Golgi
Cis face (Convex): where proteins
enter. Vesicles leave from trans
Vesicle coat proteins: COP II (ER to
Golgi), Clathrin (Golgi to cell memb),
COP I (Golgi to organelles)
Packages proteins, adds prosthetic
groups, glycolysation/sulfation.
Transfer vesicles: btw ER & Golgi.
Secretory: about to leave cell.
Present in cells with a lot of RER.



Cell Organelles 2
Mitochondia
Outer membrane (porins), inner membrane (ETC) cristae (folds), matrix (Krebs,
own DNA, calcium granules)
Steroid-secreting cells have tubular cristae
Lysosomes
Digest macromolecules, pathogens, old organelles
1 (pre-ingestion, uniform appearance), 2 (post, mottled) and residual bodies
(products not expelled from cell)
Cytosol is basic in case of leaks
Peroxisomes (microbodies)
Lighter than lysosomes, contain crystals
Peroxide & lipid metab.
Inclusions (no membranes)
Lipid droplets: In adipocytes (signet rings).
Oil Red O dyes lipids, but are usually washed out
Stores lipids and fat-soluble vitamins
Adrenals, gonads, sebaceous, liver
Melanocytes
Melanocytes in upper layer of dermis, deposit melanin there.
Block UV, reduce visual noise
Reduced amounts in substantia nigra of Parkinsons brains
Glycogen granules
Liver cells, muscles, neutrophils



Cytoskeleton Summary
Helps cell change shape, moves organelles
Microfilaments (5 nm)
Actin: broken down by profilin/thymosin, repolymerized by formin/ARP
Myosin: pulls actin
Sits on terminal web (stability)
Movement; cell adhesion (viniculin links actin w/ integrin receptors); shape changes (cytokinesis)
Found in motile cells
Toxin: cytochalasin (stops repolymerization)
Intermidiate filaments (10 nm)
Scaffolding
Formed like rope (alpha helix monomers form sheets of tetramers, which coil around each other)
Keratin in epithelial, vimentin in connective, desmin muscle,
Microtubules (25 nm)
Alpha and beta monomers. Form protofilaments (13 protofilaments per microtubule)
Centriole/basal bodies: 9 + 0 triplets
Centriole surrounded by satellite bodies (gamma monomers), bind to microtubules (form organizing center)
- ends of microtubules face centriole
2 centrioles make up mitotic spindle, attached to kinetochore of chromosomes w/ aster microtubules
Cilia/flagella: 9 + 2 doublets (axoneme in center)
o subunit has 13 subunits, 10. Doublets joined by nexin.
Cilia: anchored in basal bodies, flagella in centriole.
Kinesin: vesicles out. Dynein: vesicles in, moves flagella
Enzyme Kinematics
Summary
Enzymes
Class I: Oxidoreductases
Oxidases, oxygenases, catalases,
dehydrogenases (move 2 H or e-)
Class II: Transferases
Kinases
Class III: Hydrolases (hydrolysis of
C-O, C-N, C-S, O-P bonds)
Peptidases
Class IV: Lyases (add/remove
water, ammonia, CO2 to double
bonds)
Class V: Ligases
Class Vi: Isomerases (mutases)
Specific activity: number of enzymatic
sites per mg protein. Reaches equil. in
pure samples.
MM assumptions: E + S ES
E + P.
Under high [S], all of enzyme
becomes ES

MM equation
V = Vmax[S]/Km+[S]
Km = Vmax/2
Inhibition
Competitive: Vmax unchanged
(can be overcome w/ a lot of
substrate)
Noncompetitive: Km unchanged
(binds near binding site, destroys
enzyme activity)
Uncompetitive: Km and Vm
decrease
Remember: Enzymes lower the
activation NRG
Stabilize transition state via
substrate strain
Entropic NRG: doesnt do work,
but rather sets up the rxn.
Glycolysis Summary
Overview:
In cytosol
Glucose 2 pyruvate
Net 4 ATP & 2 NADH
Key steps:
Hexokinase, PFK, and pyruvate kinase have
free NRG, drive rxns forward.
PFK is committed step
Fates of Pyruvate
Krebs Cycle
Lactose & NADH (anaerobic & RBCs) lactose
dehydrogenase
Alanine (-CH3 becomes NH3)
2-3 BPG shunt
1,3 BPG -- (2,3 BPG mutase) 2,3 BPG.
Bypasses ATP-producing step
Lower heme affinity for O2 in low oxygen
environs
Hormonal regulation
Insulin stimulates glycolysis (increases F 2,6 bi
P) AND lowers blood sugar (cells take it in)
GLUT 2: liver/pancreas; GLUT4: fat, muscles;
GLUT5: small intestines/liver. All go to cell
membrane
Glucagon inhibits glycolysis, inc
gluconeogenesis
Inhibition
G3P dehydro inhibited by arsenate and
iodoacetate

Steps (w/ enzymes)
Glucose -- (Hex) -- Glucose 6P-- (p-
glucoisomerase) -- Fructose 6P -- (PFK) --
Fructose 1,6 bi-P -- (F bi-P aldolase) --
dihydoxyacetone P & glyceraldehyde 3-P
[interconvert due to triose isomerase] -- (G 3P
dehydrogenase) -- NADH & 1,3 BPG --
(phosphoglycerate kinase) -- ATP & 3
phosphoglycerate -- (PG mutase) -- 2 PG --
(Enolase) -- phosphoenolpyruvate -- (Pyruvate
Kinase) -- ATP & Pyruvate
Pyruvate kinase / PG kinase are misnomers!!!
1 ATP used at Hex. Step and 1 at PFK
Regulation of enzymes
Hex inhibited by its product; Glucokinase
(isomer of hex) has a high Km for glucose, not
inhibited by product. Works best at high glu
conc. (such as liver portal vein)
PFK: inhibited by ATP, citrate, and low pH
(from lactic acid). Activated by AMP and
fructose 2,6 bi-P
PFK2 makes this F 2,6 bi-P from FGP. More ATP,
PFK2 is phophpho, less active
Pyruvate Kinase: inhibited by ATP,
phosphorylation, and alanine; activated by
Fructose 1,6 bi-P
Read clinical correlations!
RNA Summary
Single-stranded, has ribose.
Replication and transcription in nucleus,
translation in cytosol
rRNA: Large & small subunit, 80% of all
RNA. Proteins + rRNA = ribosome
tRNA: clover leaf. Anticodon, CCA-3
acceptor region for a.a. Made by RNA pol
III.
mRNA: codons identical to the sense
(coding) DNA strand, but have U instead
of T.
Prokaryoytes- mRNA polycistronic (1
strand = many genes), have Shine-
Delg Sequence for ribosome binding.
Eukaryotes- 5 cap: protects from RNA
exonucleases, binds to ribosome to
start translation
RNA pol/transcription
3 to 5, error-prone (RNA temporary),
no primer needed, eukary has 3 RNA
pols
On DNA: +1 = start of transcription,
not translation.
Promoters: need to be by start site.
Enhancers: not position-dep, make
transxn faster (Eukary only)


Post-trans processing
rRNA: split in two, always = amounts due to
tandem repeated arrays, methylated,
becomes ribosome. Happens in nucleolus.
tRNA: introns removed, methylated,
glycolated
mRNA: Cap & 3 poly(A) tail added, introns
removed, exons spliced when spliceosome
[snRNA + proteins] recognizes 5 GU, 3
AG, A branch site in middle to removes
lariat. Splicing inc. protein diversity
Inhibitors
Rifampicin (some TB resistant): inhibits
beta subunit of prokary RNA pol
Amanitin: Blocks RNA pol II
Acintomyocin: stops DNA from unwinding
Ethidium Br: intercalcates btw bases, stops
unwinding
Other
Pro-opiomelanocortin is eukaryotic
poltycistronic gene
Mature onset diabetes of young:
glucokinase gene spliced incorrectly in
pancreas, not insulin dependent
Krebs Summary
Basics
Pyruvate loses CO
2
to become acetyl CoA.
(pyruvate dehydro). NADH also made.
PDH has 3 enzymes (PDH, dihydrolipyl
transferase, dihydrolipyl dehydro), 5
cofactors (thiamine pyrophosphate, FAD,
Lipoic acid, NAD, CoA)
Pyruvate
Regulation
PDH: Acetyl CoA, NADH inhibit. ATP
favors phospho (inactive) form. Insulin
favors dephospho (active).
ATP inhibits citrate synthase; ATP/NADH
inhibits isocitrate dehydro; all above + GTP
inhibits oketoglu dehydro; malonic acid
inhibits succinate dehydro; fluoroacetate
inhibits aconitase
Also limited by availability of acetyl-CoA,
activity of ETC.
PDH irreversibly damaged by arsenite
Deficiencies
Often cause brain/developmental defects
Steps (w/enzymes)
Acetyl CoA + oxaloacetate --(citrate synthase)--
citrate-- (aconitase) -- isocitrate -- (isocitrate
dehydo) --NADH + CO
2
+ oketoglutarate --
(oketoglu dehydro + CoA) --NADH + CO
2
+
succinyl CoA -- (succinyl CoA synthetase) --GTP +
succinate -- (succinate dehydro) -- FADH
2
+
fumerate --(fumerase) -- maltate-- (maltate
dehydro) -- NADH + oxaloacetate
TCA intermediates in biosynthesis
Citrate: fatty acid, sterols
oketo: glutamine, glutarate
Succinyl CoA: heme
Oxalo: carbs, nucleotide, asp/arg
Anaplerosis
Balance. Puts more acetyl-CoA in TCA by
breaking down stored fats/sugars (like btw meals),
pulls intermediates away from biosynth rxns.

Electron Chain Summary
Overview
Inner mitochondrial membrane
Electrons pump H+ into mitosol, form pH
gradient
O2: final e- acceptor, makes water
ATP synthase: powered by H+ falling
down gradient. 3 binding sites for
ATP/ADP + Pi (chemiosmotic
hypothesis)
Shuttles
Because inner memb sel perm, need
shuttles to oxidize NADH and bring it
into matrix to be reduced again
Malate-Asp: Brings NAD+ inside
More of this in LIVER and HEART: more
NRG
oglycerol-phos: FADH2 donates e-
directly to UbiQ pool, but bypasses
proton pump
Tricarboxylate: Brings acetyl-CoA inside
by turning it to citrate (only when Krebs
cycle is slow ) for lipid biosynthesis
NADH pumps 3 H+; FADH2 only pumps
2 H+



Steps (w/ complexes)
NADH to UbiQ ox-redase (Complex I):
pumps H+
[FADH2 to UbiQ (Complex II)]
UbiQ to CytochromeC ox-redase (Complex
III): pumps H+
CytC oxidase to O2 (Complex IV): pumps
H+
UbiQ: long/non-polar, found in membrane
Complexes have Fe-S clusters or
cytochromes (like heme w/ Fe or Cu)
Inhibitors
Uncouplers: allow H+ to diffuse through
membrane instead of ATP synthase, makes
heat
2,4 dintrophenol, pentachlorol phenol,
thermogenin (good for babies)
Valinomycin: transport K+ instead of H+,
disrupts pH gradient
Oligomycin: inhibits ATPase
Rotenone/Amytal: Complex I
Antmycin A: Complex III
CN & Azide: Complex IV (enters cell
rapidly)

Nucleus/Nucleolus
Summary
Bringing proteins inside:
Importins (alpha and beta): recognize NLS
(Nuc. Local. Signal) on protein of interest,
bind it. This complex is recognized by the
nupe, passes through pore.
RanGDP (in pore) helps NLS pass
through. Releases INSIDE of nucleus
when it is replaced by GTP. Process done
by G-exchange factor, called RCC1.
Importins recycled: pass back through
pore w/ help of RanGTP (alpha importin
needs CAS also). GTPase activating
proteins help re-form RanGDP, and
importins are released OUTSIDE of
nucleus into cytoplasm.
CAS re-enters nucleus through diffusion.
Cytoplasm: GDP -(pore)GTP : Nuc.
Protein released inside.
Nuc: GTP(pore)GDP : Cytoplasm
Process can begin again
Nuclear Lamina
Maintain integrity of cytoskeleton and bind
chromatin (intermediate filaments)
Lamin binding proteins: Nurim, Emerim, & MAN1.
Lamin A and B are made from same gene, spliced
differently.
Lamin C: Spliced differently, has no CaaX box (for
binding). Uses its N-terminus to bind to Lamins A
and B to form a scaffold.
LAP: Lamin binding site (to chromatin)
Dystrophies and cancer: caused in part by nuclei
w/out laminar support
Cajal: make snRNP and snoRNP (small nucleolar
ribosomal nuc prot). Found near nucleus.
Nucleolus:
Pars Fibrosa: where RNA is being transcribed
from DNA
Pars Granulosa: proteins associated w/ modifying
RNA
rRNA subunits are split and spliced, except 5S.


Transport Across Membranes
Nernst Eqn
E
ion
(mV) = 60/z *log C
out
/C
in

Na is high on outside of cell (E ~ 60), K is low on outside of cell (E ~ -90)
Total of solutes on inside of cell is ~300mM. Always assume this.
Cholesterol: too much = rigid membrane. Too little = fluid
Carbs: sialic acid residues (on membrane). Neg charge. Flu virus binds to it.
Ficks Law: if you have a thin membrane w/ a lot of surface area, your solutes will
diffuse through at fast rate.
Diffusion coefficient when temp and viscosity and molecular size .
Osmosis: net diffusion of water
Osmolarity (Osm):
60 mM of glucose = 60 mOsm while 60mM NaCl =120 mOsm (salt dissociates)
Hyperosmotic soln: cell shrinks as water leaves (more solute/less water outside) (greater osmotic pressure
outside). Opposite for hypoosmotic soln
Tonicity: concerned w/ NaCl ONLY.
Hypertonic: water leaves, cell shrinks. Hypotonic soln: little or no salt (<300 mOsm or 150 mM salt). Cell
swells. Isotonic ~150 to 155 mM salt.
Cell Volume Regulation
RVD: anything that allows K to leave cell (K-channels, K/Na symporters). Water leaves, cell volume falls.
RVI: allows Na to enter cell
NaCl: impermeable. Urea: permeable.


Mitosis/Cell Proliferation
5 phases:
Go (senescence)
G1 (GAP1) make proteins for DNA
replication
Checkpoint 1: excision DNA repair
S phase - DNA synthesis
G2 (GAP2) make proteins for cell division
Checkpoint 2: mismatch DNA repair
M (mitosis) - division of the nuclear material
Cell cycle controlled by 6 proteins:
Cyclins: regulate CDK activity
D (G1 phase) , E (Chkpt 1), A (S phase),
B (G2/M).
Cyclin dependent kinases (CDKs): protein
kinases (4 in G1, 2 in middle phases, 1 in G2)
Cyclin dependent kinase inhibitors (CDKIs):
inhibit CDK (INKs)
Wee kinase: inhibits CDKs by phospho Thr
14 /Tyr 15. Expressed at Checkpoint 2 so any
errors in DNA replication are fixed before cell
continues.
Cdc25c phosphatase: activates CDKs by
dephospho. Thr 14 /Tyr 15
CDK-activating kinase (CAK): activates CDKs
by phospho. Thr 172 /Thr 161
***Cyclins/CDK complex provide specificity for
which proteins need to be made in each phase.

Before Mitosis:
E2F (regulated/sequestered by Rb):
transcription factor needed. W/out Rb, E2F
too active, proliferation goes unchecked.
Causes retinoblastoma. CDK4
phopsphorylates Rb, which activates E2F,
which in turn makes Cylcin E & CDK2.
INK4 (p16): inhibit Cyclin D only. Cip/K
inhibits all cyclins. Expressed at checkpoints
ONLY if there is a problem during
transcription. p53 makes these inhibitors.
Phosphoryltation causes lamins to dissasmble
during prophase, reappear in telophase
Cancer Correlations
ATM (ataxia-teleng gene): Stops cell
proliferation to check for errors at Chkpt 1
Tumor supressing genes: p53, p21, p27.
Checks for DNA errors. If too many, p53
levels stay high, tells cell to undergo
apoptosis.
Cyclins/CDK can act as oncogenes
Protein-Mediated Transport & Ion Distribution
Important Factors
Rxn rate: The more solutes that bind, the
faster the transport rate.
Saturation: Transport maximum = Tm
Specificity: if it transports an L-isomer, it
wont transport the d-isomer
Comp. inhib.: many monosaccharides
compete w/ glucose transport.
***Low Km = high affinity = high
transport rate.
Channels (most common!)
Aquaporins = water. Pore diameter too
small for solute mlx. Mainly in RBC and
epithelia lining kidney, lung, intestines.
Gap jxns: smooth & cardiac muscles,
epithelia/endothelia. Non-selective
pores.
Ion channels: Voltage-gated (Na+ gate),
ligand-binding (conformational change
opens it), and mechanical distortion
(pressure)
Carriers
Only open at one end
Binding site hidden on the side of
membrane where cargo is released.

Pumps
Primary: uses ATP to pump against
conc/elec gradient.
Na/K ATPase: Most common pump! o and |
subunits. 3 Na out, 2 K in. Controls cell
volume and resting potential.
Inhibited by ouabain (thus Na cant leave cell.
Affects Na/Ca exchanger, and more Ca
available inside heart muscle to restart heart
contractions).
Ca
2+
ATPase: in muscle. Sends Ca out
Secondary: using NRG from ion falling
down its conc/elec gradient to push
another ion up its gradient. Na is usually
the driver solute.
Include cotransporters and
countertransporters (like Na/glucose cotrans
or Na/Ca countertrans.)
Ions Movement (in/out of cell)
Youll get a + Nernst potential if conc. grad.
for a cation (like Na) is high outside cell (due
to their movement into cell).
Gibbs-Donnan: Cell interior has
impermeable, mainly neg proteins. Could
attract outside cations and cause swelling if it
werent for pumps.
Translation Summary
Ribosomes
Prokaryotic ribosomal subunits: 50S + 30S
Mitochondria: 12S + 16S
Eukary: 60S + 40S
E, P, A sites: tRNA binds to A site, peptide bond forms at
P, released at E.
tRNA
Made in nucleus
Needs amino acyl in order to bind codons.
iMet tRNA (fMet in prokary) can only recognize start
codon (AUG), and not internal AUG. It binds initiation
factor. Mmet binds elongation factors.
Always read codons and anticodons in 5 to 3 direction.
Stop codons: UGA, UAG, UAA
Prokaryotes
Shine-Delgarno sequence (AGGAGGU) is always in front
of a start codon (Eukaryotes have a Kozak region)
Mitochondria
Import most translation proteins from host
Stop codons for mitochondria (AGG and AGA)
Inhibition
Prokary: Buy AT 30 (subunit), CELL at 50
A = Aminoglycosides (Streptomycin, Neomycin)
T = Tetracycline
C = Chloramphenical
E = Erythromycin
Eukary
Diptheria toxin: inhibits elongation factor 2
Cyclohexamide: 80S
Ricin: 60S

Initiation (Eukaryotes)
eIF4E binds 5 cap, eIF4B phosphorylates it so it can
release cap. eI4G is scaffold.
5 UTR stop sign always in front of AUG, needs to
be unwound w/ elF4A helicase. Initiation factor 4F.
iMet tRNA needs eIF2-GTP for activation/recognition
of start codon AUG. eIF2-GDP needs a GEF
(guanine exchange factor) to become GTP. THIS
STEP CAN BE REGULATED!
met tRNA/eIF2 combines w/ 40 S subunit to become
43S Preinit Complex.
eIF4 + 43S Preinit Complex = 48S Preinit Complex.
Goes to AUG start codon, but needs 60S subunit to
start translation!
When 60S binds (w/ eIF5), 80S subunit complete
OVERVIEW:
[eIF4(E, G, A)] + [eIF2 + tRNA + 40S] + 60S
Termination of translation
Eukary: eIF6 (dissociation factor) makes 80S leave
mRNA
Prokaryotes: 3 release factors. 1 is a GTPase
Eukaryotes: 2 release factors. 1 is a GTPase
Iron
Fe levels in cell low: need more transferrin receptors.
Transferrin RNA is not degraded. If too high: ferritin
RNA is not degraded, more toxic Fe can be bound by
newly-made ferritin.

Embryology (Week 1 and 2)
Week 1 - Fertilization
Sperm passes corona radiata, binds to ZP3
receptors on zona pellucida.
Acrosome enzyme on sperm head released
(digests ZP)
Fast block: ion channels on egg open up
(depolarization)
Slow block: cortical granules on egg
membrane (oolemna) release enzymes that
cause the ZP to harden (cortical rxn)
Primary oocyte: caught in prophase II of
meiosis until sperm enters
Pronuclei from parents meet, complete meiotic
divisions. Sperm centrioles used here.
Cleavages form morulla.
Morulla becomes blastocyst: outer trophoblast
surround inner cell mass. Uterine fluid fills
blastocoel.
Blasto leaves Fallopian tube for uterus (takes
5 days). Safe from toxins.
ZP degrades, blasto hatches and is sticky.
Blasto attaches to uterus w/ integrin.
Trophoblast cells invade uterine wall
(syncytiotrophoblast). Begins to differntiate!
Trophoblast secretes HCG hormone

Week 2 Blastocyst
Trophoblast Differentiation:
Cytotrophoblast: forms chorionic villi.
Syncytio.: becomes placenta. Dissolve
nearby capillaries in uterus to nourish
blasto.
Inner Cell Mass (2 Layers):
Epiblast (upper layer; forms amnionic
cavity).
Hypoblast (lower layer; forms primitive
yolk sac). AKA exocoelomic membrane
Mesoderm:
Forms btw hypo & epiblast.
Extraembryonic coelem (used to be
the blastocoel) forms in mesoderm,
splitting it into sphlanchnic (only
around yolk sac) & somatic (around
embryo and coelem) layers.
Primary yolk sac pinches off of larger
secondary yolk, stays beneath embryo.
Prechordal Plate:
AKA buccopharyngeal membrane
Thickening of the hypoblast. Becomes
mouth.
Primitive streak formed in epi layer.
Embryology (Week 3)
Week 3- Gastrula (3 layers)
Primitive pit: in front of primitive
streak. Cells of epiblast enter the pit,
form a neural groove (notochord).
Remember tissue demo in class!
The 1
st
cells displace the hypoblast
and become the endoderm. The next
cells to enter become the mesoderm.
The cells on top become the
ectoderm.
Gut tube forms in the endoderm
above the yolk sac.
Neural crest (on side of neural tube)
forms in the ectoderm, induced by
notochord.
Mesoderm forms paraxial,
intermediate, and lateral plates.
2
nd
Week
3
rd
Week
Cell Signaling Summary
Overview
Endocrine: signal goes through blood
Paracrine: acts on nearby cells
Autocrine: acts on self
Signaling by cell-membrane proteins
Lipid Ligands (hydrophobic)
Have a long-lasting effect.
Bind to transcription factors (either in
nucleus or cytosol) which then bind to
DNA. Their binding removes inhibitory
proteins.
All lipid receptors have similar DNA-
binding domains (hormone response
elements)
Need carrier proteins to transport them
through the cytosol to nucleus.
Ex: Steroids, thyroid hormones
Protein Ligands (hydrophilic)
Bind to surface receptors, activate
secondary messengers, have a quick
physiological response.
Stored in vesicles for later use.
Ex: Insulin

2 messengers
cAMP
Ligands bind to surface G-proteins (o subunit
inactive w/ GDP, becomes active w/ GTP).
Ligand-receptor complex causes this change.
o w/ GTP activates effector (adenylate
cyclase), which makes cAMP.
Adenylate cyclase inhibited/stimulated by
Gi/Gs proteins.
Deactivated by phosphodiesterase.
cGMP
Made by guanylyl cyclase.
Nitrous Oxide also stimulates cytosolic
guanylyl cyclase.
Protein kinase activity
Activated by cAMP
Goes on to phospho other proteins
Calcium & phosphoinositides
Gq proteins stimulate phosphoino.
Makes IP3 (releases Ca ions) and DAG
(activates Protein Kinase C).
2 messengers are always present in the cell




Epithelial Summary
Background
Forms linings
Cells very tightly packed
Layers
Simple: one layer
Stratified: 2+ layers
Look at top layer to determine cell type or see if
theres keratin
Pseudostratified:
Different heights. All cells touch basement memb.
Often have cilia
Shape
Squamous: flat
Cuboidal
Columnar
Transitional
Is stratified, but change shape. Found in bladder,
ureter.
Glands (lined by epithelial cells)
Exocrine: Connected to duct
Endocrine: No duct, relies on nearby capillaries
Merocrine: release only product (sweat)
Apocrine: some cell parts released (mammary)
Holocrine: entire cell destroyed in release (sebaceous)
Mucous secretions: contain glycoproteins
Serous secretions: proteins in water
Seromucous: in btw.
Regions
Apical
Cilia (motile, have microtubules, sit on
basal bodies, have a raggedy
appearance).
Microvilli (increase surface area, sit on
terminal web, have actin, have sticky
glycocalyx, have uniform/brush-like
appearance).
Lateral
Zona occulada: forms seal, keeps
material from seeping btw cells
Zona adherens: Made of actin. Used in
adhesion & to withstand mechanical
stress.
Gap Jxns: used in communication, have
pores (connexons).
Desmosome: Button shaped, have
keratin, dont encircle cell.
Hemidesmosomes sit on cell bottom.
Basal
Basal lamina: secreted by epithelial
cells. Contain collagen & laminin.
Basement memb: Basal lamina +
reticular lamina (made by connective
cells)
Differentiation in Embryo
Background
Embryonic stem cells transcribe our entire genome at a very low rate.
Terminally-differentiated cells arent replaced after they die (neurons and muscle)
Muscles only grow in size: hypertophy. Cancer = hyperplasia.
Induction: occurs in gastrulation when different cell types come in contact.
Notochord induces neural crest formation.
frog neural fold transplantation experiment
Ectoderm derivatives
Makes neural crest, epidermis, gut lining
Neural crest makes connective tissue of the face, peripheral nerves, APUD cells
(make bioactive amines), and melanocytes
Mesoderm derivatives
Paraxial (somites)
1 somite per body segment. Sclerotome = vertebrae. Dermatome = dermis.
Myotome = skeletal muscles.
Intermediate (urogenital)
Lateral (axial skeleton)
Endoderm derivatives
Pinching of aminion helps form gut tube. Part of yolk sac gets drawn up into it.
Endoderm forms lining of digestive and respiratory system.



Birth Defects
Stages
Gametogenesis
Fertilization
Germ disk forms
Implantation
Embryonic period (1
st
8 weeks). Very vulnerable to birth defects.
Any problems here will cause death of fetus
Fetal period
Any problems here may not result in death, but may affect how well fetus grows (intrauterine growth retardation)
Causes of birth defects
Genetics (main cause)
Infections
Drugs
Sample from amniotic fluid (or chorionic villi) is grown on culture & made into karyotype to find chromosomal
abnormalities.
Autosomal Chromosome Problems
Trisomy 21: Down's Syndrome
Trisomy 18: Edward's Syndrome
Trisomy 13: Patu
Rocking-chair foot
Sex Chromosomes Problems
Result from incorrect meiotic divisions (causes monosomy and trisomy)
Turner's: mentally normal (XO)
Kleinfelter's: XXY, XXXY

End of Exam 1
Connective Tissue
Background
Maintains form and structure
Stores fat
Defense against pathogens
Come from mesenchyme cells
Made of cells, ground substance, and fibers.
Fibers
Collagen
Most abundant protein
Provides strength/flexibility
1/3 glycine. Also has hydroxyproline and
hydroxylysine
Made of polypeptide chains (o-chains) which
form triple helix (done w/ help of registration
peptides)
Leaves cell, removal of reg peptides =
tropcollogen (type I common). Forms fiber
bundles.
Banding pattern on fibers: 64 nm
Elastic fibers
Stretches and relaxes easily. Very resistant to
degradation.
Made of elastin (desmosine and isodesmosine)
and microbrils (fibrillin protein) Microfibrils
surround elastin.
Branches (UNLIKE collagen)
Reticular fibers
Heavily glycosylated Type-III collagen.
Forms a hammock for cells




Ground Substance
Clear gel
Fills spaces
Lubricant
Barrier- very viscous
Glycosaminoglycan (GAG)
Made from repeating Uronic acid + Hexosamine sugars.
Most sulfated, attracts water.
Proteoglycan
GAG w/ protein core attached (resembles pipe cleaner)
Glycoprotein
Protein is main component, has branched carbs.
Helps in adhesion
Found in basal lamina
Cells
Fixed: Makes the fibers and ground substsance.
Fibroblasts, adipocytes, chrondocytes, osteocytes,
reticular cells.
Wandering: Made elsewhere, transported by blood to
the matrix.
Macrophage: contain lysosomes
Plasma cell: has clock face nuclei, makes
antibodies
Mast cell: histamine/heparin granules



Connective, contd
Connective Tissue Proper:
Dense Regular: resists one-
directional pulling forces.
Collagen fibers run in same
direction. In tendons.
Dense Irregular: Collagen w/out
direction. Can resist pulling
forces in multiple directions.
Surrounds organs.
Loose (areolar): Mostly ground
substance. Open to infection:
has macrophages. Not as
strong as regular, but more
flexible. Dermis, gland/cavity
linings.
Specialized CT
Elastic CT: has elastic fibers,
yellowish. Rare.
Reticular CT: very little ground
substance, looks like a giant net.
Mucous CT: mostly ground
substance (hyaluronic acid). Few
fibers, some fibroblasts
Adipose CT: has reticular fibers.
Mostly cells (unilocular and
multilocular). Highly vascularized.
Also cartilage, bone, blood.
Diseases
Marfans: fibrilin gene mutation.
Tall, disproportionate skeleton.
Ehlers-Danlos: hyper-stretchy skin.
Collagen fibrils uneven in diameter,
uneven spacing btw them.

Cartilage
Background
Avascular
Bears stress w/out being deformed.
Shock absorber
Structure
Cells
Come from progenitor (mesenchyme) cells
Chrondoblasts: active. Found in perichondrium.
Chrondocytes: surrounded by matrix. Live in
lacunae.
Isogenous group: cluster of recently divided
cells.
Ground substance
Proteoglycan (aggrecan) & glycoprotein
(chrondronectin & fibronectin)
Territorial matrix: immediately around cell. More
GAG, fewer collagen. Interterritorial matrix:
farther from cell, lighter staining.
Fibers
Mainly type II collagen
Hyaline Cartilage
Most common
Type-II collagen
Fetal skeleton
Articular cartilage = joints. No perichondrium
Trachea, larynx, plates of growing bones
Fibrocartilage
Lots of Type I and II collagen
Btw. dense CT and hyaline cartilage.
No perichondrium
Vertebral discs (annulus fibrosous)
Elastic Cartilage
Yellowish
More elastic fibers.
Type-II collagen
Ear, epiglottis
Joints
Synarthroses: limited/no movement
Synostosis: bone to bone (skull)
Synchondrosis: bone to hyaline cartilage
(ephyseal plates in bones)
Syndesmosis: bone to dense CT (fibula-tibia)
Diarthrosis: free movement, most common
Capsule: fibrous layer (dense CT) and synovial
membrane (lines articular cavity)
Ends of bones surrounded by capsule
Synovial membrane A cells: phagocytic, clear
debris . B cells: make fluid.
Problems
Osteoarthritis: loss of articular cartilage
Rheumatoid arthritis: inflammation of synovial
membrane, cavity calcifies.


Bone
STRUCTURE
Cells
Osteoblasts: secrete osteoid
Cuboidal, lots of RER, found on bone surface.
NOT epithelial tissue!
Osteocytes: surrounded by matrix
Dont make new matrix. Cant divide.
Have filopodial process: extension of cell.
Joined by canaliculi: extension of lacuna.
Osteoclast: macrophage that breaks down matrix.
Contains acids. sits in Howship capsule. Has
ruffled edge to increase surface area. Many
nuclei.
Periosteum: Surrounds bone
Blood supply & source of osteoblasts
Fibrous layer: Dense CT (collagen/fibrovlasts)
Osteogenic layer: cellular, bound to bone by
Sharpeys Fibers.
Endosteum: Lines interior of bone
Thin, reticular fibers
Also has osteoprogenitor cells
Matrix (fiber, ground substance, minerals)
Osteoid: Type I collagen, ground substance, no
mineral. Helps give it shape, resistance to tension.
Mineral: rigid, but not flexible. Calcium, phosphate
(make hydroxyapatite).
Calcification: need osteocalcin, osteonectin,
phosphotase enzymes.

Calcium mobilization
Parathyroid Hormone
Acts on osteoblasts. Blasts stimulate osteoclasts,
which activity. Bone breaks down, frees Ca
(resorption)
Calcitonin
Acts on osteoclasts. activity. Bone formation.
Spongy Bone
Primary or secondary bone
Location: epiphyses (long bone ends), interior, center of
flat skull bones
Forms trabeculae (bone spicules) projecting into
cavities for marrow
Osteocytes in spicules dont form neat lameallae.
Compact Bone
Secondary bone only!
Location: right beneath periosteom and in diaphyses
(long bone middle).
Forms osteons (Haversian sysytems)
Haversian canal: center. Holds nerve, blood vesses, lymph.
Runs along length of bone.
Volkmanns canal: Connects Haversian canal
Collagen fibers in same lamellae are parallel, but crisscross
fibers in adjacent layers.
Osteon surrounded by cement (minearls w/ few fibers).
Lamallae farthest from canal formed 1
st

Outer circum. lamallae: Beneath periosteum. No
osteons.
Inner circum. lamallae: Surround marrow. No osteons.
Intersitial lamallae: Remains of old osteons

Bone, contd
TIME OF SYNTHESIS
Primary Bone
Always spongy
Collagen fibers more unorganized, less
minerals.
Temporary, except in tooth sockets & flat
bone sutures.
Secondary Bone
Spongy or compact
Collagen fibers more organized
Mature bone
Osteons continuously remodeled
As bone grows, marrow cavity expands.
Compact bone thickness stays same.
Hormones
Growth Hormone: stimulates chrondocytes
Vitamin A: stimulates osteoblasts
Vitamin D: Calcium absorption
Estrogen: stimulates bone formation
METHODS OF FORMATION
Intramembraneous Ossification:
Formation of skull bones (derived from
neural crest)
No cartilage model formed. Mesenchyme
goes straight to osteoblasts
Endochrondal Ossification:
Formation of long bones.
Mesenchyme forms chrondocytes, which form
epiphyseal plates (responsible for growth)
ZONES:
Proliferation: isogenous groups stacked one on top of
the other as they divide.
Hypertrophy/Calcification: chrondocytes start
mineralizing, blood vessels can enter.
Ossification: cartilage is now bone.
Cartilage begins in long bone center. Pushes outwards
to both ends during growth.
Ossification Centers:
1: Middle of long bones
2: In epiphysis. Form after primary.
3: Form tubercles and ridges.
Fracture Repair
Blood vessels break. Clot forms around fractured ends.
Progenitor cells enter, become osteoblasts,
chronodocytes, fibroblasts.
Callus formed (CT and cartilage model). Replaced by 1
bone, then 2 bone.
DISORDERS
Dwarfism: epiphyeal plates understimulated
Chrondodysplasia: Type II cartilage mutation = short
deformed limbs
Osteogenesis Imperfecta: Type II cartilage mutation = short,
fragile limbs
Rickets: Ca deficiency
Osteomalacia: osteoid doesnt mineralize = soft bones
See Clinical Correlations for more!
Gluconeogenesis & Glycogen
Background
Glycolysis in reverse. 6 ADP equiv, 2 NAD+ made.
Diff enzymes used for irreversible rxns
Primary location: liver. Makes glucose for brain, RBC
NRG (ATP) comes from fatty acid oxidation!
ATP, citrate, and acetyl CoA stimulate gluconeo
Key Steps (w/ enzymes)
Pyruvate - (pyruvate carboxylase) oxaloacetate
(PEP carboxykinase) PEP
Reverse of Pyruvate Kinase
Pyruvate carb uses ATP, biotin, K, Mg to add CO
2

to pyruvate.
Oxaloacetate leaves mitocho via mal-asp shuttle.
PEPCK is in cytosol.
PEPCK uses GTP, K, Mn to remove CO
2
, add Pi
If ATP low, oxalo goes to Krebs.
Fructose 1,6 BP (Fructose 1,6 phosphatase)
Fructose 6P
Reverse of PFK1
RATE-LIMITING STEP!
Glucose 6P (Glucose 6 phosphatase) Glucose
Reverse of hexokinase
Membrane-bound in ER
No net flow of carbons from acetyl CoA to glucose.
Most sugars, most aa (not lys & leu), succinyl-CoA and o-
keto are glucogenic
Glucagon stimulates gluconeo, but slows glycogen
synthesis. Phosphorylates synthase & phosphorylase.
Insulin inhibits gluconeo & stimulates glycogen synthesis


Non-pyruvate sources
RBC and muscles make lactate from pyruvate when O
2
low.
Liver removes extra lactate by using it for gluconeo (Cori
Cycle)
NADH made from breaking down EtOH. Too much NADH
inhibits rxns that need NAD+. Also, fatty acid oxidation in liver
and gluconeo from lactate is inhibited.
Alanine (pyruvate + NH
2
) also fed into gluconeo. Makes urea.
Glycogen Synthesis
Glucose (hex) Glu 6P (phosoglucose mutase)Glu 1P
(G1P uridyl transferase)- UDP -glu
Add UDP-glucose to protein glycogenin (done by transferase
for 1
st
2 residues). Glycogenin primes itself up to 8 glucose
residues.
Additional glucoses added by glycogen synthase. Forms 1,4
glycosidic linkage on non-reducing ends. Stimulated by glu-6P
Once chain has ~11 residues, branching enzyme removes 6
residues, forms 1-6 linkage w/ nearby chain.
Regulation
Dephospho form of glycogen synthase (a) is active and is
NOT sensitive to stimulation by glucose 6P
Phospho form (b) is inactive and sensitive to stimulation by
glucose 6P
Several kinases and 2 messengers aid in regulation
Glycogenolysis
Glycogen phosphorylase breaks 1-4 linkage in glycogen,
makes glucose 1P. Regulated by phophorylase kinase.
Phospho form = a = active.
Debranching enzyme when you hit last 4 glucoses of each
chain, it puts 3 on the C4 of next branch (last hydrolyzed by
water).
Glucose 1P later becomes glucose 6P, enters glycolysis.



Connective Tissue Fibers
Collagen
Background:
Abundant (~30% of all proteins)
Gly every 3
rd
aa. Forms tighter turns than ohelix. ochains
form triple helix.
Type I: Ligaments, tendons, skin
Type II: Cartilage
Type III: Skin vessels, tendons
Type IV: Basement membrane
Type V: Fetal membranes
Collagen Synthesis (w/ enzymes)
In RER lumen : some pro & lys get hydroxylated by prolyl
& lsyl hydroxylases. Cofactor: ascorbic acid (vitamin C).
Lys later glycosylated (glycosyl tranferase). Helps form
preprocollagen left-handed triple helix. Signal peptide
cleaved in RER, becomes procollagen.
Helix secreted. Peptidases cleave registration peptides on
ends. Procollagen becomes tropocollagen. Forms
microfibrlis, then fibers.
Lysyl oxidase: deaminases chain and turns it to an
aldehyde so it can undergo crosslink rxn. w/ other
microfibrils. Cofactor: copper.
Lysinonorleucine: main collagen crosslink.
If not properly made, fibrers gets degraded.
Elastin
Background:
Not a triple helix. Gly not every 3
rd
residue.
Mostly nonpolar aa. No carbs!
Only one type.
Desmosine: main crosslink. 4 aa join together, forms
heterocyclic crosslink.

Proteoglycans
Lubricates, supports. Antithrombic (heparin)
GAGs: long chains of repeating disaccharides.
Proteoglycans = Mucopolysaccharides = GAGs
bound to core protein
GAG Structures:
Heparin: GlcUA or IdUA (Sulf) + GlcNAc (Sulf)
Heparine sulfate: same as above, but fewer S
residues.
Dermatan sulfate: GlcUA or IdUA + GalNAc (S)
Chrondroitin sulfate: GlcUA + GalNAc(S)
Keratan sulfate: GlcNAc (S) + Gal
Hyaluronic acid: GlcUA + GlcNAc
Adhesive Fibers
Help in cell movement, adhesion, wound healing.
Fibronectin: fibroblast, endothelial cells, monocytes,
neutophils
Has binding domains that bind proteoglycans,
collagen, fibrin, integrin (in cell membrane).
Helps cells detect changes in extracellular
matrix.
Chrondonectin: cartilage
Lamin: epithelial cells
Also has different binding domains.
CT Fibers, contd
Disorders:
Scurvy: w/out vitamin C, collagen not hydroxylated. Get weak chains.
Crosslink rxn inhibited by sulfhydryl groups (found in homocystine)
Lathyrism: -aminoproprionitrile (from sweet peas) inactivates lysyl oxidase.
Decreased crosslinking of collagen. Also caused by Penicillamine (used in copper
chelation therapy).
Collagenase: In tissue- reshapes collagen. In Clostridium histolyticum (bacteria)-
causes gangrene (bacteria breaks down fibers to spread).
Elastase: Breaks down elastin.
Antiproteases: oantitrypsin normally in balance w/ elastase, except in
emphysema (too moch elastase).
Hurler: No a-L-iduronidase. Cant make dermantan, heparin S.
Hunter: No a-L-iduronidase-2-sulftase. Same as above.
Sanfillippo A-D: No heparin made. See clinical correlations for enzymes affected.
Rheumatoid arthritis: Hyaluronic acid depolymerized.

Pentose Phosphate Pathway & Other Sugars
Pent Phos Path (2 Phases)
Makes NADPH for biosynth (H+ donor, not carrier)
Makes nucleotides from Rib 5P
Not very active in brain/muscle tissue
Oxidative Phase (non-reversible)
Glu-6P (Glu-6P Dehydro) NADPH + 6P glucono-
lactone (Lactonase) 6P-gluconate (6P-Gluco
Dehydro) NADPH + CO
2
+ Ribulose 5P
Non-Ox Phase (reversible)
Rib 5P + Xyl 5P (transketolase) G 3P +
Sedoheptulose 7P - (transaldolase) Fru 6P +
Erythrose 4P
Ery 4P + Xyl 5P (transketolase) Fru 6P + G 3P
Phosphopentose isomerase & epimerase turn Rib 5P to
Xyl 5P
Transketolase needs thiamine!
Net: 3 5C

sugars turned to 2 6C & 1 3C sugars
Regulation
Fru 6P & G 3P can be diverted from glycolysis if Rib 5P
(but not NADPH) needed.
Can become Glu-6P (gluconeo) if more NADPH needed,
Glutathione and Anemia
Ox. Glutathione + NADPH (Glut Reductase) Red.
Glut + NADP+
-SH on glutathione helps RBC scavenge free radicals in
lungs. Otherwise, oxidative damage causes hemolytic
anemia.
Glu-6P Dehydro deficiency: Cant make red. glut w/out
NADPH. ox stress, sensitivity to drugs that cause cell
protein ox, sensitivity to fava beans (have pro-
oxidants).



Very common inborn metabolic defect. Helps combat P.
falciparum malaria, which needs lots of reduced glut in
RBC.
Metabolizing Non-Glucose Sugars
Maltose, sucrose, lactose dissachs broken down to
glucose, fructose, galactose.
Fructose Metabolism
Enters cells through GLUT-5. Sperm fuel.
Fruc-(Fructokinase)-Fruc-1P-(Aldolase B)- DHAP+ D
glyceraldehyde (triose kinase) G 3P
G3P enters glycolysis. Bypasses insulin and PFK1
regulation. Kinase uses ATP.
Fruc not metabolized as well as glu.
Fruc Intolerance: W/out Aldolase B, Fruc-1P
accumulates. Traps available Pi, inhibits ATP synthesis
Galactose Metabolism
Gal (Lactokinase)- Gal 1P + UDP-glu
(Uridyltransferase)- UDP-gal + Glu 1P
Galactosemia: W/out lactokinase or UD transferase, Gal
1P can become galacticol, accumulates in eye. Pi also .
Sugar Alcohols
Glu- (aldolase reductase)-sorbitol-(sorbitol dehydro)-Fruc
Diabetics have more glu, more aldolase reductase is
induced. Sorbitol forms crystals which build up in eyes,
kidneys.
Glucuronic Pathway
Uses glucoronic acid to make GAG (proteoglycans),
glycoproteins, glycolipids
UDP-glu (UDP-glu dehydro) UDP glucoronic acid.
Most animals can make Vitamin C from glucoronic acid.
Humans cant.
Resting Membrane Potential
Background
Cells resting membrane potential is overall negative.
Goldman-Hodgson Eqn.
V
m
= 61 - log P
K
[K
+
]
o
+ P
Na
[Na
+
]
o

P
K
[K
+
]
i
+ P
Na
[Na
+
]
I

Nernst for more than 1 ion.
Cl- already near cell resting potential, ignore its contribution to eqn.
P = permeability constants
The more permeable a membrane is to an ion, the closer the resting membrane potential will be to
that ions Nernst equilibrium
Ohms Law
I
ion
= g
ion
(E
membrane potential
E
Nernst
)
Current =I, conductance = g
If the membrane potential equals the Nernst potential for an ion, youll get 0! Therell be no current!
Terms
Hyperpolarized: membrane potential is MORE negative relative to outside.
Less excitable. RMP.
Depolarized: membrane potential is LESS negative relative to outside.
More excitable. RMP . Approaches threshold!
It always takes the cell awhile to respond to a stimulus. Not instantaneous!
Cell is 100x MORE permeable to K than to Na, so Na will always slightly hyper/depolarize cell.

Action Potentials & Propagation
Electrotonic Potentials:
No threshold (soft & hard touches detected); passive
conductance; size-graded; not self-generating; doesnt
propagate; no refractory period; spatial/temporal
summation
Mechanoreceptors (skin), chemoreceptors (taste buds)
Action Potentials:
Threshold potential; all-or-nothing conductance; stimuli all
produce same amplitude; self generating; can propagate;
refractory period needs to end before channel can re-
open; potentials arent summed.
Nerve and muscle cells
Phases of Action Potential
During threshold and rising phase, membrane becomes
more permeable to Na. Voltage-gated Na channels open.
Cell interior becomes more positive.
Peak: 1
st
Na channels to open start to close their
inactivation gates (right before peak). Activation gates
stay open until voltage falls. K channels begin opening, K
leaves cell.
During repolarization phase and at rest, membrane is
most permeable to K. Na channels closed.
Afterhyperpolarization: K channels start to close. Resting
membrane potential restored.
Voltage-Gated Channel Details
Na channel: 2 gates. Active state (both gates open),
inactive state (inactivation gate closed, cant conduct) and
resting (active gate closed, ready to be opened by
stimulus)
K channel: Only one gate that opens at threshold.
Absolute Refractory Period: From start to point of highest K
conductance. Na channels in inactive state, cant be opened.

Relative Refractory Period: After Peak. Na channels at rest and
ready to open again, but some K channels still open. Stimulus
needs to be much higher than threshold voltage to open them
(accomodation).
Toxins
Tetrodotoxin (TTX ): Blocks voltage-gated Na channels.
Tetraethylammonium (TEA): Blocks voltage gated K
channels
Propagating Action Potentials
Time Constant: t = R
membrane
C
membrane
Capacitance = membrane ability to store chage (rate of
charging up). Low capacitance means faster propagation.
Resistance: higer in small diameter nerves (ions hit each
other rather than flow through cytosol).
Refers to time it takes nerve to reach 63% of threshold.
R = 1 / g (conductance)
Length (Space) Constant: = \(R
memb
d / R
interior
4)
Non-myleinated nerve diameter would need to be 4X as
large to have same propagation speed as myleinated ones.
R
memb
needs to be high to keep ions from leaking out.
Space constant is distance it takes for nerve A.P. to lose
63% of its original voltage.
Non-Myleinated Nerves
Na influx leaves net charge outside membrane, which
attracts neighboring Na ions, and they leave a net behind,
and so on.
Myleinated Nerves
Saltatory propagation: action potentials travel to Nodes
Schwann cells: insulation, increase memb resistance, lower
capacitance. Cell charges up faster, ions travel down axon
farther.
Blood Clotting
Background
Involves: 1) Constriction of injured vessel 2) Clumping of
platelets to plug hole 3) Clot formation on top of plug
Platelets
No nuclei, disc-shaped, contain granules.
Activated by collagen: become spiky, release granules.
Adhere to each other and collagen w/ von Willebrand
Factor.
Platelet Granules
Thromboxane A
2:
: stimulates ADP release
ADP: promotes platelet aggregation
Vasoactive amines (norepinephrine, serotonin):
constrict inured vessel.
Fibrinogen
Has 2 , 2 , 2 subunits. Joined by disulfide linkages
(knots).
Thrombin cleaves off part of and chains, leaves most
of fibrinogen intact.
Cleaved chains on top of fibrinogen are electostatically
repulsive, but once removed fibrin fibers (monomers)
can aggregate.
Fibrin covalently joined, but flowing blood could disrupt it.
Needs to be stabilized by transglutamase (Factor 13).
Activated by thrombin.
Thrombin
Inactive form in blood = prothrombin.
Factor 10 a (w/ 5a and Ca++) cleaves 2 places btw
Arg/Thr and Arg/Ile. Forms 2 chain enzyme joined by
disulfide bond. B-chain is trypsin-like (cleaves Arg-Gly)
Thrombin makes fibrin monomers, and activates factors
5, 7, 8, 13.

Clotting Factors
10a: activated 2 ways:
Extrinsic = Tissue Factor (Factor 3) from wound
forms complex w/ 7a and Ca. Complex activates 10.
7 activated by wounded tissue or thrombin.
Contact w/ glass & kallikrein activates 7.
Intrinsic: Factor 12 in blood activated by wounded
tissue. 12a 11a -9a -8a 10a. Cascade!
Redundancy: if no 12, Tissue Factor activates 11. 7a
can also enter intrinsic by activating 9.
Modification of factors
Before activation, they are proproteins. Before that,
they are pre-proproteins (still have signal sequence).
Carboxylation: allows Ca to bind. Factors 2, 7, 9, 10.
Glu aa becomes carboxylglutamyl (gla) at N-
terminus.
Vitamin K dependent carboxylase makes gla.
Vitamin K1: comes from plants.
Gut flora: make MK-n (menonquinone). Has more
double bonds.
Factor 9 has Gla residue. 11a cleaves it twice, but 9
is still docked to membrane.
B/c of gla, thrombin (w/ Ca) can bind to phospholipid
membrane of platelets rapidly. Increase cascade by
10,000x!
Prothrombin has NO gla residues (needs to move
around in blood).
W/out K, you cant clot quickly!

Blood Clotting (Part II)
Clotting Regulation
Coumarins (anticoagulants):
Vitamin K epoxide is reduced to its quinone form by
Vit-K reductase. This enzyme is inhibited by the
coumarin Warfarin.Thus, clotting factors arent
carboxylated!
Overdose of coumarin can be fixed w/ dose of Vit-K.
Antithrombin III: forms irreversible complexes w/ thrombin
& activated factors, inhibiting them.
Heparin: helps antithrombin III bind to factors quickly.
Used as a fast anticoagulant during surgery.
Protein C: inactivates 8a and 5a. Activated by
thrombomodulin-bound thrombin, which is found on lining
of healed blood vessels.
Protein Z: binds w/ protease inhibitor to inactivate 10.
Antitrypsin: inhibits proteases, including many of the
clotting factors
Lipoprotein associated coagulation inhibitor: inhibits
Tissue Factor-10a complex.
Clot Dissolving
Plasmin: Protease made in kidneys. Breaks up fibrin.
TPA: tissue plasminogen activator (cleaves plasminogen
into plasmin). In wounded tissue.
PAI: plasminogen activator inhibitor.
Urokinase: removes clots in kidney.
Streptokinase: used mainly in European hospitals.
Complexes w/ plasminogen to activate it.
Treatments
Asprin: inhibits formaation of thromboxane A2
Plavix: inhibits ADP and platelet aggregation

Diseases / Disorders
Abnormal Clotting (genetic):
Antithrombin 3 deficiency
Protein C deficiency: causes venous thrombosis
Protein S deficiency: C cofactor
Factor 5 lieden: 5 cant bind Protein C = excess
clotting.
Factor 20210a: makes prothrombin mRNA very
stable, and more prothrombin made than can be
broken down.
Excess fibrinogen
Clots can travel through blood, cause stroke,
heart attack, pulmonary embolism.
Oral contraceptives increase this risk by decreasing
antithrombin 3)
Dietary:
Christmas Dinner: Factor 7a increased by high
lipid meals, can cause clots.
Abnormal Bleeding (genetic):
VKCFD: not carboxylating enough of your clotting
factors.
Von Willebrand Disease: poor platelet adhesion
Hemophilia: mostly factor 8 mutations.
Some hemophiliacs received blood transfusions from
people w/ HIV in the early days.
Post-clotting
Bradykinin dilates healed blood vessel, increases
its permeability. Cleaved from kininogen by
kallikrein. Must be remade constantly: 10% lost in
lungs.




Blood
Composition
55% Plasma, 40-50% RBS, ~1% WBC
Plasma
90% water. Albumin protein maintains osmotic pressure.
Carries metabolic waste. Yellow from iron
RBC
Have no nuclei! But do have mitochondria (make heme),
cytoplasmic enzymes.
Membranes have carb compobebts that determine blood
type.
Amount regulated by hypoxia, erythropoietin (from
kidneys), filtration by spleen.
Reticulocytes: Young RBC which still have RNA. Slightly
more purple. Presence indicates patient trying to produce
more RBC.
PROBLEMS:
Microcytosis: too small thassalemia, Fe deficiency
Macrocytosis: too big Vitamin B12/Folic acid deficiency.
Hyperchromia: too dark more Hb b/c cell is small
Hypochromia: too pale Fe deficiency
Spur cells: spiny shape - renal problems
Basophilic stippling: Pb binding to heme indtead of iron,
cell trying to make more Hb.
Howell Jolly Body: nucleated RBC not removed by spleen
Platelets
Come from megakaryocytes in marrow.
Form plugs by binding to basement membrane of wounded
epithelial cells.
No nuclei, but have mitochondria & glycogen
Overproliferation: means body is stressed.

Leukocytes
Granulocytes
Neutrophils: neutral colored granules.
Poly-lobed nuclei.
Younger forms have U-shaped band nuclei
Engulf and digest bacteria using activated O
2

35-62% of all WBC. Granulocytosis= large amount
present (during infection). Neutropenia = low amounts
(during leukemia treatments)
Eosinophils: Red granules
Bi-lobed.
Large granules w/ histamine, peroxidases, crystals.
Crystals have bar-shaped major basic proteins.
Combat worms, but can be over stimulated during
allergic rxns.
1-4% of WBC
Basophils: Blue granules.
Grabules have heparin, histamine. Granule contents
released after IgE binding.
<1% of WBC
Mononucleocytes (agranular)
Monocytes: very few granules
Called macrophages when they leave blood & enter
tissues (osteoclasts in bone)
3-8% of WBC
Lymphocytes: blue cytoplasm
B-cells: humoral immunity. Make a lot of antibodies.
Include plasma cells, memory cells.
T-cells: cellular immunity. Enhance other WBC (helper
CD4), inhbits (CD8), or destroy cells on contact (killer)
20-45% WBC

Hematopoeisis
In fetus
Yolk sac (makes angioblasts and vessles); liver
(platelets, granulocytes); spleen (RBC); T-cells (thymus)
Bone Marrow
Marrow in axial skeleton makes blood throughout
adulthood. Long bones stop after adolesence.
Biggest cells: adipocytes & megakaryocytes.
Blood made on hematopoietic cords. Stroma:
connecctive tissue. Sinusoids: capillary system that
allows cells to enter main circulation.
3 developing WBC to every 1 developing RBC b/c RBC
live much longer, dont need to make as many of them.
Marrow is also where Fe recycled from heme.
Early Cells
Colony-forming cells: young cells that can form a colony
of hematopoietic cells. Come from multipotent stem cells
Stimulated by cytokines (growth factors). G-CSF =
makes granulocytes. M-CSF = monocytes. Interleukin-5
= eosinophils. GM-CSF / thrombopoietin = megakary.
GM-CSF / IL5 = RBC
Cytokines used in clinical settings. However, early ones
tend to cause leaky capillaries.
RBC Formation
Proerthroblast: dark blue cytoplasm, large nucleus
Basophil erythroblast: blue cyroplasm, smaller nucleus
Polychroamtic erythriblast: checkerboard nucleus
Orthochromic normoblast: small nucleus, pinkish
cytoplasm
Reticulocyte: no nucleus
RBC

Granulocyte Formation
Myleoblast: large nucleus, medium blue cytoplasm
Promyelocyte: 1 granules overlie nucleus
Myelocyte: named for type of granules it has.
Smaller nucleus, pink cytoplasm
Metamylocyte: Mardi-gras mask nulceus
Band form: U-shaped nucleus
Mature granulocyte.
Monocyte Formation
Monoblast
Promonocyte: may have folded nucleus
Monocyte / macrophage
Other Cells
Lymphoblast lymphocyte
Megakaryoblast megakaryocyte-platelets
Diseases
Deficient synthesis
too little marrow, cytopenia (too few of a cell
type), aplastic / Fanconis anemia (too few cells
in general)
Disordered hematopoeisis
Granulocytes may have too many nuclear lobes
Macrophages may not break down ingested material
(Gauchers Disease)
Overproliferation
Leukemia
Gene Regulation
Diff. btw Prokary & Eukary
Eukary DNA wound up tightly in chromatin, while bacterial
DNA quickly accessed
Bacteria: 1 RNA Pol. Eukary: 3 RNA Pols.
Eukary RNA undergoes modifications after transcription
Eukary RNA must pass through nucleus to be translated.
In prokary transcxn & translxn are coupled.
Prokaryotic Regulation
Depends on initiation, termination, and stability of mRNA
(which is rapidly degraded)
Operon: has control sequences that are associated w/
structural gene sequences.
Catabolic operon: when a certain nutrient is present
(inducer), operon begins transcxn of genes to metabolize
it. Ex: Lac operon (breaks lactose to glu)
Anabolic operon: w/out an essential nutrient, operon
remains on in order to synthesize it. Ex: Trp operon (trp is
co-repressor, turns operon off if too much Trp made)
Features of Operon
Promoter: where RNAPol binds. Upstream of init site.
Operator: where repressor protein binds
Regulator: codes for repressor protein. Always transcribed.
Not always in same operon.
cAMP receptor/catabolite activator protein (CRP/CAP):
helps RNApol bind to DNA w/out falling off. By promoter.
Attenuator: terminates transcxn if an RNApol accidentally
slips by repressor.
Lac Operon Regulation
Codes for 3 genes, most important is Lac Y (permease:
brings things into cell)
W/out lactose, transcription continues at very
low level (that way, some permease is always
present to bring in lac if it appears).
Repressor: coded by Lac I gene. Tetramer:
together the subunits bind strongly to DNA.
Inducer is a negative allosteric modifer, and
when it binds to repressor it causes it to release
DNA
Repressor recognizes palindrome sequence of
operator (RNApol does not, since its
unidirectional)
Catabolite repression: if glucose present
(preferred food), Lac operon turned off (Cap
levels drop b/c less cAMP made)
Eukaryotic Regulation
Cis-acting: gene sequences that regulate the
same gene. Need to work w/ trans-acting.
Trans-acting: other elements that act on the
gene (like repressor or RNApol). Bind to cis
CTCF can enhance or inhibit transcxn (can
block RNApol, keep chromatin from unwinding,
etc). Its zinc fingers can bind to DNA in diff ways
depend on what fxn is needed. It can bind
insulators.
Silencer: binds repressor (eukary version of
operator)
Gene Regulation II
Eukaryotic Regulation, contd
DNA-binding Proteins
Protein surface complementary to surface of double helix.
Subunits each have weak hold, but together bind very
strongly. Subunits can be joined by:
Homeobox: have helix-turn-helix domain that holds
DNA (ex: CAP)
Zinc Finger
Leucine Zipper
Antisense RNA
Insulin-like growth factor receptor 2: Both alleles have 2
promoters.
From mom, downstream one is methylated. Can make a
sense RNA that makes Igf2. Always transcribed. From dad,
upstream one is methylated. Antisense RNA is made, which
helps to down-regulate this gene.
If you inherit 2 copies from one parent it can be fatal. This
is one reason why humans arent parthenogenic.
SiRNA/microRNA
siRNA becomes double-stranded so that dicer protein can
recognize / cut it. Becomes single strand again, binds to
antisense mRNA (w/ RISC protein). RISC degrades target
mRNA by slicing. Humans use it to attack HIV RNA.
miRNA down-regulates mRNA by forming UTR-stem loops
Riboswitch RNA
Riboswitch RNA folds up gene-coding sequence into an
inactive form.
When a certain protein or chemical binds, RNA unfolds into
an active form and translation of gene sequence occurs
faster.






Alternate Promoters
Dystrophin gene: longest human gene. Has 8
promoters w/ their own exons. Each one makes a
tissue-specific product. More variety in mRNAs
produced by alternative splicing, adding poly-A
tails, etc.
Cancer
Overall, cells very good at removing spontaneous
mutations. Most cancers caused by 3-7
independent changes in cell regulatory genes
(unless youre already born w/ one)
Oncogenes: need only one mutant allele (loss of
heterogeneity) to cause uncontrolled proliferation
Tumor-suppressor genes: need both alleles to be
mutated to cause loss of normal fxn. Rarer.
Loss of p53 fxn found in ~50% of all Stage IV
carcinomas.
Mdm2 overproliferation binds too much p53, keeps
it from initiating transcription of regulatory genes.
(Ex: it may not be able to make apoptotic proteins
to destroy faulty cells)
Li Fraumeni syndrome: inherited mutant p53.

Cell Junctions
Basic Info
Tight Jxns (zonula occludens)
Forms blood-brain barrier in brain capillary linings.
Zonula Adherens
Desmisomes (Macula Adherens)
Gap Jxns
Proteins
Tight Jxn: transmembrane proteins (occludin & claudin); actin
Na/K pumps in retina have reversed polarity: found in apical membrane instead of
basolateral
Zonula Adherens
Calcium-dependent adherin jxn (cadherins); Actin
E-cadherin: found in epithelail
N-cadherin: found in neurons.
Desmosome
Desmosomal cadherins, keratin
Hemidesmosomes have integrin. Keratin
Gap Jxn
Connexin proteins (~20 types): 6 per connexon
Close to prevent toxic amounts of Ca or H+ from spreading to neighboring cells.
Disease
Pemiphigus: autoimmune desmosomal disease of skin proteins

Myoglobin & Hemoglobin
Hemoglobin
4 chains, each w/ O
2
binding site
Chains cooperatively bind O
2
: 1
st
doesnt bind easily, but makes it easier for successive O
2
to bind
Readily releases in low oxygen environments (tissues). Binding sites only fully saturated in lungs.
and chains prevalent post-birth. chain in fetus, releases O
2
from placenta.
OxyHb looks red. DeoxyHb looks more blue. Hb bound to CO looks bright red.
R-form: relaxed (high affinity for O
2
). T-form: tight (deoxy Hb).
Myoglobin
1 chain w/ single O
2
binding site
Accepts O
2
from Hb in muscle
Heme
Porphyrin: attached to F chain (by proximal histidine) and E chain (distal histidine)
Fe needs to be +2 state, or else it will bind but not release O
2

CO & CO
2

80% flows in blood as bicarbonate. 20% bound to Hb (carbamate), but not at O
2
binding sites.
CO does bind to O
2
binding sites. Hb reverts to R state, and holds on to remaining O
2
rather than
release them.
Bohr Effect
Hb readily releases O
2
in low pH environments (tissue w/ more lactic acid)
2,3 BPG
- charges on BPG stabilize the + charged cavity on T-form Hb (deoxy). Causes Hb to release O
2
more
readily.
Sensory Receptors & Transduction
Background
Sensory cells convert physical stimuli (mechanical,
chemical, light, heat) to electrical signals.
***Nerve cells are influenced by neurotransmitters
across a synapse!
Adequate stimulus: lowest stimulus picked up by a
receptor.
Receptor (generator) potential: caused by stimulus,
sets off action potential if it reaches threshold. Na
doesnt influx through voltage gated channel.
Made up of 1
st
order (receptor) neuron, which synapses
w/ at least 2 other neurons
More action potentials can be generated based on the
intensity and duration of stimulus.
Classes of Sensory Receptors
Exteroceptors: stimuli come from our senses
Interoceptors: concerned w/ internal environment of
bodies.
Proprioceptors: relay info about our bodies in space.
Mechanoreceptors
Meissners Corpuscles: fingertips
Hair Follicle Receptors: detect movement on skin.
Merkels Disks: non-hairy skin
Tactile Disks: similar to above
Ruffinis Corpuscles : detect stretching in joints
Pacinian Corpuscles: Sub Q layers of non-hairy skin,
muscles:
- Contain nerve ending wrapped around gel & conn tissue
- Initial pressure causes nerve fiber to deform. Gel allows
return to shape. When pressure removed, nerve deforms
again (intensity change)
Other Somatic Sensory Receptors
Thermoreceptors: 36-45 C = warm
Nociceptors: respond to anything that will cause tissue
damage
Photoreceptors: Na gate open in the dark, closes in
light (activated by hyperpolarization)
Chemoreceptors
Sensory Attributes
Modality: specific tissue receptors and neuron
pathways for each stimulus: light only detected by eye
pathway (labled line).
Intensity:
Frequency code: More a.p. for more intense
stimuli
Population code: More a.p. for more receptors in
the region
Duration: nerves can adapt rapidly (phasic) or slowly
(tonic) to a continuous stimulus
Location:
2 point threshold: ability to discern this depends
on amount of nerves in region getting poked by 2
needles.
Lateral inhibitory region: surrounds excitatory
region so that we can localize where a stimulus is.
Somatosensory Pathways
Dorsal column: 1
st
order neuron on left side of body;
2
nd
order neuron crosses over to right side of the body
in brain stem. Pressure, proprioception.
Anteriolateral: 2
nd
order neuron crosses over to right
side in spinal cord. Temperature, pain.



Gene Technology
Cloning
Making exact copy of DNA sequence of interest
Making Vectors
Vectors: allow you to insert & copy any DNA sequence of
interest.
Plasmid: bacterial. Usually have antibiotic resistance
genes.
Cosmid: viral (bacteriophage)
Yeast Artificial Chromosome: Can hold thousands of
base pairs.
Restriction endonucleases: cleave palindromic
sequences, make sticky ends. Allow unrelated DNA to be
put together w/ ligase.
Techniques
Plasmid: Insert gene of interest into antibiotic resistance
gene. Bacteria now sensitive to that drug (screens).
Expression plasmid: gene of interest is expressed, fused
to bacterias normal protein products from its own
operons.
CaCl2: makes it easier for bacteria to import recombinant
plasmids. Neutralizes DNA charge so it can enter
membrane.
cDNA
cDNA: made from mRNA using reverse transcriptase.
cDNA now only contains exon sequences for your
plasmid (bacteria cant remove introns)
cDNA collection = cDNA library. When placed into vectors
= expression library.
Genomic library = entire genome (exons & introns).
Probes & Hybridization
Probe: finds a specific DNA sequence in cDNA library
(antibodies find expressed proteins in expression
library)
Hybridization: when probe binds to its complimentary
DNA strand. Can bind strongly to one sequence
(stringent), and less strongly to others.
Southern Blot: seeing which sequence probe binds too.
Run DNA through gel (smallest pieces move farther).
Blot it from gel onto filter paper so its more durable.
Probe will hybridize to target sequence on filter paper.
Western Blot: Proteins. Northern Blot: mRNA
You can use probes to see if a patient has a mutant
allele for a given DNA sequence. Mutations will give
you a different length RFLP from normal.
Removal of tandem repeats in a mutant allele is
another way to see what disease a patient has.
Sequencing (Building Probe)
Use deoxynucleotides to build a chain. Use
dideoxynucleotides to stop elongation
PCR
Rapidly amplifies DNA sequence.
Use taq polymerase (from thermophilic bacteria): can
withstand heat.
Practical Uses
Transgenic animals: mixture of self genes and genes
from other stem cells inserted into them.
Pharming
Making an ovum express the DNA of sequence. The
adult will produce in large quantities a protein product
that it normally wouldnt make (like a drug in its milk)


Synaptic Transmission
Electrical Synapses
Ions move through hemichannels (connexons in gap jxns)
Bidirectional flow of ions
Connexons only 5nm apart
In smooth & cardiac muscle
Chemical Synapses
Most common type of synapse
Synaptic cleft = 30-50 nm
Unidirectional flow of transmitter
A.p. in pre-synaptic cell opens Ca gates, which enters
presyn nerve, promotes release of neurotrans vesicles.
Release neurotrans, which bind to ligand-gated non-
specific channels (both Na & K move through). This
binding acts as generator potential, which sets off action
potential (opens fast Na channels, and later K channels).
Channel closes when neurotrans unbinds.
Excitatory (open Na channels) & inhibitory (open K or Cl)
post-synaptic potentials affect the action potential.
SLOWER than electrical synapses.
Neurotransmitter Details
Quanta: the release of products from a vesicle. Vesicles
hold same number of neurotrans molecules (quantum).
Need to release a lot b/c some neurotrans diffuse away or
get degraded (Ach degraded by Acetylcholinesterase)
Synaptic Contacts
Axosomatic: axon synapses to soma. Strong signal,
greater influence on trigger zone
Axodendritic: synapses to shaft/spine. Signal decays if its
too far from soma, so you need a lot on dendrites
Axo-axonic: synapses to axon. Inhibitory or excitatory for
neurotrans release (affects Ca influx). Doesnt affect
signal.
Input/Output Relationships
One-on-One: single a.p. in presynaptic cell to 1 a.p. in post
synaptic cell. Neuromuscular jxn
One-to-Many (Divergence): 1 a.p. to many a.p. in many
post synaptic cells. Renshaw cells in spinal cord
Many-to-One (Convergence): many a.p. to 1 a.p. Most
common one. Motor neurons
Summation & Other Occurences
Spatial: when 2 separate presynaptic signals reach post-
syn nerve at same time
Temporal: when same pre-syn nerve sends 2 consecutive
signals to post-syn nerve
Pre-syn Inhibition: Block Ca channels, lessens Ca influx
into presynaptic axon, fewer transmitters released to bind
post-syn. No a.p. made. Done at axo-axonic synapse.
Post-syn Facilitation: More Ca flows in. TEA can also do
this.
Potentiation (rapid firing of pre-syn): Some residual Ca remains
in pre-syn before it can diffuse out. Several effects:
Facilitation: More neurotrans released, then falls off quickly
(short-term memory)
Posttotonic: Neurotrans falls off slowly (long-term memory)
Depression: depletes all neurotrans vesicles (why seizure
end)
Drugs
Nifedipine, nisoldipine (dihydropyridines): Ca antagonists.
Block Ca channels. Decrease blood pressure!
Neurotransmitters
Neurotransmitter Criteria
Has to be made in neuron
Released by pre-syn neuron to affect post-syn
neuron
Exogenous application of substance mimics
endogenous release
A mechanism exists for removing it from cleft.
Types
Small molecule (fast acting).
Sensory signals to brain, motor signals to
muscles
Large molecule (slow acting): cause more prolonged
response
Long term opening or closing of channels
Classical Neurotransmitters (small mlx)
Acetylcholine
Released by cholinergic nerves. Made in
mitochondria from acetyl CoA & choline.
Used in neuromuscular jxns. Inhibitory for
cardiac muscle.
2 molecules bind to -chains of receptor
Catecholamines (Biogenic amines)
Epinepherine, Norepinepherine, Dopamine
Deficiencies = bipolar disorder, depression
Made from tyrosine. Dopa hydroxylase turns
DA into norepi.
Broken down by monoamine oxidase.
Pargyline (antidepressant) blocks MAO (norepi
breakdown), more stays in cleft. Imipramine
(better antidepressant) blocks norepi reuptake.

Reserpine blocks storage in vesicles so it cant be
released (tranquilizer)
Dengeration of dopaminergic neurons associated w/
Parkinsons. Treated w/ L-Dopa.
Seritonin (5-HT)
Eating, sleeping, arousal. Inhibitory effects.
Prozac blocks 5-HT reuptake only.
Glutamate
Memory, learning
Excitatory
GABA (a and b receptors)
Inhibitory. Stabilizes neuron electrical activity.
A: increases Cl influx. Benzodiazepine: facilitate
GABAs fxn by further calming down neron activity.
B: increases K conductance.
Glycine: inhibitory
Neuropeptides (large mlx)
Usually affect metabolism, gene acitvation in nucleus.
Include opiates (regulate pain)




Neuromuscular Junctions
Structure
1-to-1 synapse
Post-syn muscle has muscle has folds to increase surface area for ACh binding.
ACh located near top of fold. ACherase ready to break it down.
Signal Transmission
EPP (end plate potential): very large generator potential. Always large enough to trigger a.p. in
muscle fiber to contract it.
MEPP (miniature EPP): Small depolarizations occur after EPP (.4 mV). Happens when a lone vesicle
in synaptic happens to release its neurotransmitters.
Drugs
Curare (tubocurarune): nicotinic receptor antagonist. Competes w/ ACh. Inhibits EPP, keeps it from
generating a.p.
Atropine (muscarininc antagonist): has no effect, b/c no muscarinic receptors at neuromuscular jxn
Myasthenia Gravis
Autoimmune: Antibodies attack folds and ACh receptors. Synaptic cleft also grows wider, so more of
ACh diffuses away. Need anti-ACherases (to preserve ACh). Immunotherapy needed (remove
thymus in adults).
Denervation
Nerve is severed. TTX-insensitve-Na channels and more ACh receptors appear to make muscle
hyper-sensitive.
Unless muscle is re-innervated, it atrophies and is replaced by CT, fat.


Heme Synthesis & Breakdown
BACKGROUND
Heme: type of porphyrin (4 pyrole rings)
15% heme synthesis occurs in liver. Rest is made by RBC
precursors (mature RBC have no mitochon)
HEME SYNTHESIS
Glycine + Succinyl CoA -- (Aminolevulinic acid synthase)
ALA (ALA dehydratase) porphobilinogen [4 needed] (PB
deaminase)- Bilane - (uroporphyinogen I synthase / UP III
cosynthase) UP III (UP decarboxy) coproporphyrinogen
III (CP oxidase) protoporphyrinogen IX (PP oxidase)
protoporphyrin IX (ferricholase) heme
ALA synthase: In mitochondria. Cofactors: Mg and
pyridoxal 5-phos. Rate-limiting step! Inhibited by excess
heme, glucose. Stimulated by barbiturates (removes
hemes by making other porphyrins). Half life ~ 1 hr.
ALA dehydratase: has its own 2 rings. Acts as a primer
in putting 4 ALA together in porphyrin ring.
CP oxidase: back in mitochondria.
-inogen rings have acetate & propionate substituents. All A
and most P later decarboxylated to methyl & vinyl.
UP III rings : AP-AP-AP-PA. UP I: last ring is AP. Non-
functioning. UP I made when UP III cosynth defecient.
HEME BREAKDOWN
Happens mainly in spleen, also liver & bone marrow
Hb globin, which break down to a.a.
Heme now exposed, oxidized to hemin
NADPH reduces hemin 2x to make hydroxyheme-oxogenase
complex.
Complex loses Fe & CO (ring opens) Biliverdin
Biliverdin Reductase Bilirubin
Travels from spleen through blood bound to albumin.
Unconjugated bilrubin (indirect)
In liver: becomes conjugated bilirubin (water-soluble). Gets
bound to 2 glucoronate. Leaves as bile.

Direct bilirubin = water soluble. Van den Bergh rxn tests for it.
Z-protein & liagndin bind bilirubin in liver cytosol.
Hemopexin: in liver. Picks up hemin from albumin
Removal From Body
Glucoronate removed in bowel. Bilirubin duced by
microorganisms to urobilin. Excreted by kidneys (makes urine
yellow).
Further reduced to stercobilin (makes feces brown).
Glutathione and Methemoglobin
Used to remove reactive oxygen compounds in RBC
Methemeglobin (Hb Fe 3+) reduced by NADH to Hb Fe 2+.
Methemeglobin has no carrying capacity for oxygen, need to
remove it!
Fe Info
Ferritin: binds Fe for storage in liver. Regulated by Iron Response
Elements on gene.
Transferrin: carries Fe 3+ into liver cell. Also has IRE
Hemosiderin: another way to bind Fe. Amorphous.
Ferroportin: sends Fe from cells into plasma.
Hepcidin: MAJOR regulator of Fe absorption. Made in liver, but
also used by macrophages & enterocytes (intestines). Binds &
destroys ferroportin, stops them from sending excess Fe to
plasma.
Enterocvytes sloughed off after ~week, their Fe gets picked
up by ferroportin.
Macrophages keep Fe, use it to kill pathogens during
infections.
Hepcidin synthesis inhibitied by EtOH.
Hemin is most easily-absorbed source of Fe. Vitamin C also
makes Fe more easily absorbed (same as methemoglobin, but
w/out globin chain)
Hb in RBC has most of our bodies Fe. Ferritin is next largest.
Heme Synthesis & Breakdown,
Contd
DISORDERS
Jaundice: Body contains excess bilirubin. Symptom, not a disease.
Deposited in skin, eyes.
Pre-hepatic: heme breakdown (from hemolysis), indirect bilirubin in blood.
Intra-hepatic: many liver cells not working (from hepatitis), slightly more indirect bilirubin.
Post-hepatic: bilirubin excretion (bile duct obstruction), direct bilirubin.
Lab tests can detect elevated levels of LDH enzyme (from lysed RBC)
Infant-mother Rh incompatibility: infants RBC lyse. Cause jaundice. Babies put under blue light to turn skin
bilirubin into more-soluble photobilirubin.
Crieger-Najar: too much unconjugated bilirubin accumulates in liver, damaging it. Can also move to brain (causes
kernicturus).
Anemia: Fe deficiency
Can cause koilonychia (spoon nails), pallor, beefy tongue.
Sickle Cell Anemia: point mutation in -globin side chain. SS = disease, SA = carrier.
SC: 2
nd
most common, another side-chain mutation. S = Thallasemia
Hemachromatosis: too much Fe
Heriditary form: caused by mutations in HFE (hepcidin not made in high amounts, Fe from intestines picked up in
large amounts by liver).
Need to remove blood (phlebotomy)
Hb is saturated, Fe carrying capacity shrinks.
Bronze Diabetes: Causes pigmentation in skin, Fe deposited in pancreatic & muscle cells, damaging them.
Secondary Hemachromatosis: ingesting too much Fe when Fe storage is already high.
Can be caused by blood transfusions for thallasemia (get good globin, but too much Fe. Need Fe chelating).
Porphyrias: accumulation of intermediates that can only be used for heme synthesis. Porphyrins in skin cause
photosensitvity. Use hemin or glucose to stop heme synthesis (basically, induce anemia).
Accute Intermittent Porphyria: Porphobilinogen accumulates, causes red urine in light.
Porphyria cutanea tarda: UP III accumulates. Causes chest hair in women
Pb inhibits many points of heme synthesis. Pb binds ALA dehydratase, but can be treated w/ Zn (its cofactor).
Binds Ferrochelatase, needs to be removed by Pb chelating agent.


Nerve Tissue Summary
Functional Divisions
CNS: Brain, spinal cord
PNS: All other nerves
Somatic: Conscious control
Autonomic: No conscious control
Classification
Motor, Sensory, Interneuron (lie btw sensory & motor)
Structure
Bipolar
Multipolar
Pseudopolar (soma lies off of axon, but signal still
passes through it). Structure of sensory nerves.
Nerve Anatomy
Soma: has pigmented inclusions
Lipofuscin- build up w/ age
Neuromelanin- found in substantia nigra neurons.
Loss associated w/ Parkinsons.
Cytokeleton
Microtubule association proteins: MAP2 in
dendrites, MAP3 in axons.
Mylein
Schwann cells (in PNS): myleinate only one
section of one axon (w/out mylein, it can surround
several nerve fibers)
Gives white matter its color (soma = gray)
Axon Hillock: has Nissl Bodies (RER & ribosomes), but
dont extend into axon.
Vesicles
Anterograde: Moves from soma to synapse. Kinesin.
Retrograde: Moves from synapse to soma. Dynein
Motor neurons
Neuromuscular jxn: axon to fiber
Motor unit: single motor neuron synapsing to
several muscle fibers. The smaller the motor unit
(the fewer muscle fibers a nerve connects to), the
more accurate and controlled the movement we
can make.
Neuroglial Cells
Oligochrondocytes: myleination in CNS (no
Schwann cells)
Astrocytes: supports nerves & blood vessels.
Fibrous = white matter. Protoplasmic = gray matter
Microglial: small nuclei. Rare. Dormant until an
injury occurs; then act as macrophages to remove
damaged cells
Ependymal cells: ciliated. Line fluid filled ventricles.
Forms a barrier.
Peripheral Nerve Organization
Endoneurium: CT bundles myleinated & non-
myleinated nerves
Perineurium: Bunle of many endos.
Epineuria: Bundle of many peris.
Autonomic Nerves
Innervate glands, smooth muscle
Dorsal root ganglion: input from sensory nerves.
Satellite cells: support root ganglion nerves.
Sympathetic nerves found in paravertebral ganglia
running along spinal cord
Enteric nerves: Line smooth muscles of gut.
Control peristalsis




End of Exam 2
Skeletal Muscle & Contraction
LOCATION
Evenly distributed through body
Attaches to bones except for facial muscles and intrinsic
tongue muscles (connects to CT)
STRUCTURE
Arise from myotomes (somites). Grow by hypertrophy
(size increase), not hyperplasia (proliferation)
Cells: multinucleated, unbranched, striated. Have
abundant mitochondria. Sarcolemma = cell membrane
Organiztion: myofilaments (actin / myosin) in
sarcomeres. Muscle fibers organized into fascicles. Each
cell surrounded by endomysium (CT w/ fibroblasts) and
smooth sarcoplasmic reticulum.
Proteins
Actin: made up of G-actin monomers.
Tropomyosin: sits on actin.
Troponin: small protein on tropomyocin. 3 units: TnT=
attachment. TnI = blocks myosin-binding site on actin.
TnC = where Ca binds, causing tropomyocin to move so
myosin can bind actin.
Myosin: 2 heavy chains in middle, flanked on each end by
2 light chains w/ heads. Has myosin & ATP binding sites.
Tropomodulin: caps actin, prevents depolymerazation.
Titin: maintains structure of sarcomere. Connected to
myosin by MyBP-C.
Sarcomere
Surrounded by Z-lines, where actin anchors.
Has full A-band in center (where myosin & actin overlap)
I band: beside A band. Actin only.
H-zone: center of sarcomere. Myosin only. Bisected by
M-line, where myosin anchors.

Other Components:
T-tubule: invagination of membrane.
Cisternae: areas of sarc ret near t-tubule (2 cisternae
surround t-tubule. Btw A and I band in humans)
Motor Units
Group of muscle fibers innervated by single axon.
Small unit: innervates fewer muscle fibers. Muscle is
dense w/ nerves, has delicate, specific movement.
Muscle Spindles
Receptors that tell how contracted or relaxed muscles
are (proprioception). EtOH inhibits this.
Extrafusal fibers: regular muscle fibers attached to
spindle.
Intrafusal fibers: inside spindle, attached to afferent &
motor neurons
Reflex arc: contraction of muscle to return it to a normal
state. Goes through spinal cord only.
Fiber Types
Type I: large motor units. Slow oxidative, contractinng.
Red b/c have more myoglobin. Mostly posture muscles.
Type IIa: Fast oxidative. Pink.
Type IIb: Fast glycolytic. Small motor units. Large
diameters. Fast contracting. White. Moslty in limbs.
Type I slow to fatigue (marathoners)
Type II quickly fatigued (sprinters)
All types found in muscle, but in different ratios.
Muscle Contraction, Contd
CONTRACTION
Sliding Filament Mechanism
Remember neuromuscular jxns! ACh is neurotrans.
A.P. travels through sarcolemma and down T-tubule.
Depolarization causes release of Ca from sarc ret.
Ca helps make sarcomere contract: moves troponin out
of way so that myosin head can attach actin.
ATP-binding lessens myosins affinity for actin. Detaches
from actin.
ATP cleavage causes head to swing down, pulling actin
fiber (power stroke). ADP still attached.
When ADP removed, head returns to rigor state. In death,
no ATP. Muscles stay in rigor state until muscles degrade
(rigor mortis).
This cycle is repeated many times in a single contraction.
Ca actively reuptaken into sarc ret, muscle relaxes.
I-band decreases during contractions.
MUSCLE STRENGTH
Isometric: crosslinks form and tension develops, but
muscle doesnt shorten (pushing hands together).
Isotonic: lifting a constant weight (afterload).
Length-Tension Retationship
If you start from rest, your muscle can generate a lot of
tension. If the muscle is already streched out a lot, you
wont generate much more tension.
Passive: pre-existing tension on muscle before you
decide to contract it.
Active: produced by contracting muslce (forming cross-
bridges)
Optimal resting state: optimal overlap of actin & myosin
fibers. Best contractions obtained


Force-Velocity Relationship
When you have no afterload: the velocity of
contraction is fastest (holding nothing)
Velocity is very slow with very heavy weights.
Tension depends on # of crossbridges formed.
Velocity of shortening depends on how fast
crossbridges formed.
Exercise
Produces an oxygen debt of ~11.5 L that must be
replenished: 3.5 L from alactic O
2
debt (phosphagen
system & aerobic); 8 L from lactic O
2
debt
ATP & creatine: used in sprints. Needs to be
replenished.
Important Terms
Single twitch: 1 action potential
Treppe: several a.p. Causes unfused tetanus.
Tetanus: several a.p. in rapid succesion. Muscle has
sustained contraction, cant relax at all between
stimuli
Disorders
Muscular Dystrophy: X-linked disorder of dystrophin
protein (helps connect muscle membrane w/ basal
lamina).
Myostatin: normally stops myogeneis in
mesenchyme and satellite cells. Overexpression =
too much muscle.
Atrophy: muscle diameter directly correlates w/
strength. Shrinkage caused by: disuse, dennervation;
cachexia (loss of appetite, esp in cancer patients).


DNA Mutations & Repair
Mutation Definition
Any permanent change in DNA sequence.
Somatic: acquired, like cancer.
Germline: transmitted to next generation.
Sources of Damage
Exogenous (physical) or Endogenous (chemical)
UV radiation; oxygen radicals; metabolically-activated
carcinogens; base-pair modifications (cytosine
deamninates to uracil); agents that intercalcate btw bases
(EtBr); agents that form interstrand crosslinks (nitrogen
mustard)
Effects of Damage
1) Repair 2) Apoptosis 3) Mutation
Types of Mutations
Base-substitution (point)
Synonomous (silent): mutant codon still makes wild
type a.a.
Missense: a. a. substitution.
Nonsense: mutant codon is stop codon.
Frame-shift: deleting or inserting1 or 2 bp causes a.a.
chain to be very different from wildtype. 3 bp still retains
most of the original a.a. chain
Splice site: mRNA chain will be too long or short. Affects
final protein length.
Slip-strand mispairing: Skipping a base. Happens in
sequences w/ a lot of repeats.
Epigenetic changes: DNA sequence remains the same
(no insertions or deletions)
Methylation: transcription factors dont bind to
methylated cytosine.
Changes in histone residues can also affect
transcription.
Other mutations
Regulatory region found in 5 end of DNA. Mutations
here affect amount of RNA made.
3 mutations affect stability of RNA (poly A tails)



Other Forms of Variation
Single-nucleotide polymorphisms: source of variation in
genome. Usually 1 bp in length. May be problematic if they arise
within the sequence for an important gene. Can cause RFLPs if
restriction enzymes recognize them.
Expanded repeats: tandem repeats found in or near disease
genes.
Microsatellites: 2-6 bp repeats. Cancerous ones differ from
hosts own
Mutation Repair Overiew
Rate of damage > rate of repair = disease.
Mitochondria: their DNA repair is less efficient, but can better
withstand damage.
Types:
Nucleotide excision: UV rays cause adjacent thymines to
dimerize, causes lesion on helix. Excision endonuclease
removes dimer, flags excision site w/ OH. DNA pol fills in gap,
ligase closes helix.
Deficient in xeroderma pigmentosum (skin blisters in sun)
Base excision: DNA glycosylase removes base, forms abasic
gap. Endonuclease nicks sugar backbone, and DNA pol
removes the entire sugar, replaces base. Ligase closes gap.
Used by mitochondria to repair oxidative damage.
Mismatch repair : MMR gene. Removes loops formed by
mismatches
Defect in this results in a form of colorectal cancer.
Repairing Double-strand Breaks
Require use of special proteins (Ku, DNA-dependent protein
kinase)
Non-homologous: trims back both ends of break site to reform
most of sequence. Not template-directed. Error prone.
Homologous: repairs original gene using 2
nd
copy of
chromosome as template.
Diseases
Helicase defect: Blooms syndrome (chromosome breaks)
Strand break: Ataxia-telangiectasia (lymphoma) and
Fanconis anemia (leukemias).


Fatty Acid Oxidation
Background
Lipolysis: breakdown of triacylglycerols
Lipogenesis: formation of fatty acids & TG
Digestion mainly in small intestines; lipogenesis in liver
Important Enzymes
Bile salts emulsify lipids.
Lingual Lipase: from tongue, works best w/out bile
Gastric Lipase (acid stable): specific for 10-12 C fatty acid
chains. Needs Ca.
Pancreatic lipase (Steapsin): Colipase stabilizes it. W/out it,
enzyme rapidly degraded when it touches oil/water interface
of fat droplets, gets easily washed off by bile.
Leptin: acts on hypothalamus, tells body to eat less and
exercise.
Disorders: too little causes adipocytes to store more fat
= obesity
Fatty Acids
palmitic acid = C16:0 (hexadecanoic acid)
stearic acid = C18:0 (octadecanoic acid)
oleic acid = C18:1 (9) [cis-9-octadecenoic]
linoleic acid = C18:2 (9,12) [all cis 9,12-octadeca]
linolenic acid = C18:3(9,12,15) [all cis-9,12,15-octadeca]
arachidonic acid = C20:4 (5,8,11,14) [all cis5,8,11,14
eicosatetraenoic]
Linoleic & linolenic: essential fatty acids (not made in body)
Arachidonic: essential-dependent
Lipolysis Regulation
Main enzyme: Hormone sensitive lipase (a = active =
phospho).
Activated by cAMP kinase, which is activated by adenyl
cyclase.
Insulin inhibits adenyl cyclase & stimulates lipase
phosphotase = inhibits lipolysis (happens when our bodys
NRG levels are high).


End product: Free FA. These inhibit lipase and
adenyl cyclase.
FA travel to other tissues bound to albumin.
Glycerol has to leave adipocytes to be activated.
Only tissues w/ glycerol kinase can readily use it
(liver, kidney, intestines, brown fat, mammaries)
Making Acyl-CoA (pre -Ox)
RATE-LIMITING STEP!
Occurs In liver mitochondria.
Turns FA to fatty acyl-CoA (FA can bind to cell
membrane, cause damage).
Acyl-CoA synthetase (cytosol) Carnithine Palmitoyl
transferase I Translocase CPTII (in outer/inner
mitochondrial membrane)
-Oxidation
Also occurs in liver mitochondria
Acyl-CoA dehydro (makes FADH2); Enoyl-CoA
hydratase (adds H
2
O); hydroxyacyl-CoA dehydro
(makes NADH); acyltransferase [thiolase] (makes
acetyl-CoA and acyl-CoA).
FA chain gets shortend by 2 C. On the last cycle
(when chain is down to 4C), makes 2 acetyl CoAs.
Ex: Palmitic (16C) goes through 7 cycles of
oxidation.
Palmitic to CO
2
& H
2
O: 7 NADH = 21 ATP (3 per
Complex I of ETC); 7 FADH2 = 14 ATP (2 per
Complex II); 8 Acetyl CoA = (12 per Krebs). 96 ATP.
Making Acyl-CoA uses 2 ATP.
Net: Indirectly makes 131 -2 = 129 ATPs.
If starting from a triacylglycerol: Glycerol to CO
2
&
H
2
O: 22 net ATP (1 acetyl CoA + 3 NADH + 2 ATP
1 ATP [to activate glycerol])
See lecture notes for tristearic example!
Fatty Acid Oxidation, Contd
Regulating -Ox
No rate-limiting step. Mainly depends on available FA.
Malonyl CoA made from acetyl CoA. Inhibits CPTI (FA
uptake into mitochondria).
Lack of NAD+ / FAD+ inhibits -Ox.
Oxidizing Odd-Chain & Unsaturated Fats
Unsat: double-bonds need to be moved for -Ox to occur. 2
additional enzymes needed: enoyl isomerase & reductase.
Odd-chains make propionyl CoA and acetyl CoA. Needs
cofactors biotin & Vit B12.
Branched chains: -ox. Branched chain on beta end of FA
makes propionyl CoA (glucogenic) and acetyl CoA
(ketogenic). Odd chaims
Peroxisomal oxidation: similar to -Ox, but makes peroxide
in 1
st
step. Immune cells use it to kill pathogens.
When a lot of FA oxidation occurs, acetyl-CoA builds up.
Liver turns it to ketone bodies (acetoacetate & 3-
hydroxybutyrate). Other tissues turn ketone bodies to acetyl
CoA and succinate for Krebs. Needs HMG CoA Synth.
Lipid Synthesis
Needs NADPH. 3 sources:
Mainly from Pentose Phos Path.
Citrate-pyruvate Cycle: activated when you have high
NRG (ATP). Citrate moves acetyl-CoA from mitochon to
cytosol. Turns cytosolic NADH to NADPH.
Isocitrate shuttle: inhibited by lots of ATP. Citrate &
isocit accumulate, go into above pathway.
Rate-limiting step: acetyl-CoA (acetyl-CoA carboxylase)-
malonyl-CoA
Regulation: Citrate allosterically promotes formation of
active acetyl-CoA carboxylase polymers
Insulin activates lipid synth. Epi & norepi inactivate
acetyl-CoA carboxylase (inactive = phospho.)


Acyl-carrier protein: maintains high NRG bond.
Priming: acetyl-CoA gets transferred to Cys group on
carrier.
Linkage: 2 C from acetyl-CoA move to malonyl-CoA.
Gives off CO
2
.
Reduction: occurs w/ NADPH. Make a 4C carrier
protein.
Linkage & reduction repeat many times to make C16
(palmitate).
When you label the C2 on acetyl-CoA primer at the
start, it becomes the last C on end of chain. C2 gets
pushed to the end of chain.
Only use 1 acetyl CoA to make palmitate. The labeled
O on malonyl CoA is given off as CO
2
.
Use reductases to add double bonds (occur at C9 in
numans first) & elongases to make chain longer!
Diseases
Long-chain Acyl-CoA Dehydro Deficiency (C > 12).
Fatal in infants.
CPTII deficiency: cant break down fats for NRG.
Causes muscle weakness, blood in urine (renal
failure).
Similar to McArdles disease in glycogen
breakdown.
Refsums: cant breakdown highly-branched phytanic
acid (from green vegetables). Body stores it instead.
-ox. Makes dicarboxylic acids, not acetyl-CoA.
Occurs when beta-ox inhibited.
Cirrohsis: too much fat limits O
2
supply to liver cells.

Smooth Muscle
Types
Unitary: lining of various tracts (GI, resp, urinary)
1 nerve on several muscle cells. Contract
together.
Intermediate filaments: desmin only
Multiunit: tunica media of arteries, hair follicles, eye.
1 nerve over 1 muscle cell. Contract
independently of each other.
Intermediate filaments have desmin & vimentin
Both come from mesoderm
CELL STRUCTURE
Tightly-packed, spindle-shaped, pale nuclei.
No striations, few mitochondria.
Loosely organized actin filaments connected to
dense bodies (act as Z-lines). Few to no myosin
(forms as needed).
Cells in muscle often lie in several layers
perpendicular to one another.
FUNCTION
Contractions: slow. Have intrinsic rhythm.
Spikes last about 10-50 msec. Plateaus have
delayed repolarization = slow contractions. Slow
waves are self-excitatory (generate own a.p.).
Act as pacemakers, seen in interstital cells of
cajal in GI tract.
Innervated w/ autonomic nerves
Parasymp: | visceral contraction, + vessel
contraction
Symp: + visceral contraction, | vessel
contration
NO motor end plates!
Have caveolae instead of T-tubules.
No troponin in smooth muscle! Instead, Ca binds
to calmodulin (which moves tropomyosin off of
actin). Unlike troponin, it phopshporylates myosin
to activate it.
Most Ca comes from extracellular fluid. Myosin
phospho lets it bind to actin, form crossbridge.
W/out phospho, myosin can bind actin very slowly
(latch-bridge formation). Lets it contract w/out
using Ca or ATP.
Sarc ret very small, IP3 releases Ca. Voltage gate
is main route for Ca influx. Also has ligand gates,
leak channels.
Ca ATPase & Ca/Na exchanger pump Ca out,
causes relaxation.
Muscle always low-level contractile (tone). Allows
muscle to maintain constant pressure on contents
of vessels.
Problems:
Smooth muscle can form plaques in blood
vessels, blocking blood flow.
Raynauds Phenomenon: abmormal skin
coloration b/c abnormal muscle spasm. Lack of
blood to local tissues (frostbite).

Cardiac Muscle
LOCATION
Found in myocardium of heart. Btw endocardium (epithelial +
CT) & epicardium / pericardium (CT).
Myocardium actually a coiled sheet. Left venticle is double
coil = more thick. Heart contracts in wringing motion.
Derived from cardiogenic mesenchyme: heart starts in
angiogenic clusters in front of head of neural plate, later
folds under.
CELL STRUCTURE
Branched. Central nuclei. Abundant intrercellular space w/
capillaries. Lots of mitochondria.
Intercalcated disks at end of cells.
Has fascia adherens: sheet instead of ring (zonula). Has
actin. On vertical axis.
Macula adherins: has desmin.
Gap jxns: on horizontal axis.
Different cardiac cells:
Atrial cells: have fewer T-tubules (slower response to
contractile stimulus). Have natriuretic granules: promotes
urination of sodium (controls volume).
Ventricular cells: Larger, more T-tubules.
Purkinje fibers: conduct nerve impulses
T-tubules at Z-lines! Terminal cisternae only on one side of
tubule (dyad). Not a lot of sarc ret (more primitive than
skeletal).
FUNCTION
Heart cells have own intrinsic rhythm. Modulated by
autonomic nerves.
Ca from cytosol & Ca
+2
-induced Ca
+2
release from sarc ret.
Contraction: starts in artria, ends in ventricles.
Cells in SA have fastest beat (pacemaker). Sends impulse to
AV.
Bundle of His: relay impulse from AV to Purkinje.
Sympatheic: speed up heart. Preganglionic motor nerves
connect to intermediolateral cell column (Clarks column) in
thoracic spinal cord.
Parasympathetic: slows heart. Motor neurons start in medulla
of brain. Ganglia right on heart (short postganglionic nerves:
release ACh).
Contraction Phases
Ventricle:
Repolarization = dip after peak. Na & some Cl enters, K
leaves. Plateau = Ca enters (through L-type channel:
long). Also induces release of more intracellular Ca.
Hyperpolarization and resting = K leaves. 250 msec.
Bundle of His & Perkinje fibers have similar action
potential.
Atrial:
Similar to above, but no plateau. Fewer Ca channels. 150
msec.
SA node:
Upstroke due to influx of Ca (T-type channel: transient =
fast), not Na. Reaches threshold faster to act as
pacemaker. 150 msec. Also seen in AV node.
Starlings Law: the more filled w/ blood the heart is at rest, the
greater the pressure at contraction. In heart, overlap at rest
(preload) is a little shorter (so it can stretch a little more and
reach optimal when blood fills heart).
Diastole: passive. Fibers at rest. Pressure right after
contraction, ready to be filled. Systole: active. Fibers stretched
due to blood before contraction. If you overstretch, the cross-
links are less optimal, pressure produced is decreased.
Cardiac contractility depends directly on intracellular Ca.
Responsible for strength of contraction (along w/ blood vol).
Cant get tetany in heart due to long refractory period! You
have to end a.p. to start another contraction. Long a.p. in
cardiac overlaps the contraction
Damage
Reperfusion injury: happens when blood rushes back in after
clot removed. O2 free radicals causes damage.

Molecular Pathology
Genetic Defects
Single Gene Examples
Entire gene deletion (Factor 8 in hemophilia)
Partial gene deletion (frameshift). Cystic Fibrosis: loses Phe. Becker dystrophy: loses 3 BP, still
has most a.a.
Tri-nuceotide repeats. Fragile X : binding site for TF heavily methylated by repeats. No
transcription!
Insertion of one sequence into another sequence (usually done by Alu sequences).
Point mutation (single nucleotide): can destabilize mRNA if it occurs in poly-A tail.
Mitochondrial genome
MERRF: form of epilepsy.
Multi-factorial (when 2 or more genes affect phenotype)
Chromosomal (can be visualized)
Monosomy, trisomy, etc
Uniparental disomy: You get both genes from one parent, nothing from the other.
Micro-deletion of chromosomal segment (Prader-Willi)
Translocations, inversions.
FISH: detects chromodomal aberrations using flouresence.
Inherited Genetic (may occur during meiosis)
Somatic Cell (not transmitted to offspring)
Mosaics: different chromosome sets or mutations found in different tissues throughout body.
Pleiotropy (1 gene influences many phenotypic traits)

Blood Vessels
Basics
Arteries and veins have 3 layers:
Tunica Intima: innermost, by lumen. Simple squamous
endothelial, basal lamina.
Tunica Media: smooth muscle.
Tunica Adventitia: may have collagen & elastic fibers.
Arterial System
Conductance: blood has highest pressure & speed.
Has thicker tunica media to withstand pressure.
Large elastic (aorta); muscular (medium); arterioles;
metartioles. Small ones usually paired w/ venule.
Venous System
Capacitance: most blood at any given time is in the
veins.
Lacks elastic fibers. Appears collapsed due to thinner
walls
Has valves.
Large (vena cava); veins; venules.
Capillaries
Capillary bed: large surface area
Blood flows slowly here (for gas exchange)
Continuous capillaries: no pores. Tight jxns
Fenestrated capillaries: pores, w/ diaphragms
Sinusoids (discontinuous capillaries): wide lumens.
RBCs arent in single file.
Lymph
Vessels = lacteals
Carries interstitial fluid from capillsries to veins
Carries aborbed fat.
Other Features
Vaso vasorum: small vessels that bring blood to
outer layers of large vessels.
Anastomes: bypass capillary beds during shock.
Carotid bodies: chemoreceptors in carotid arteries
that measure blood O2 and pH levels.
Pericytes: lie on outside of capillaries. Can become
new vessels in case of injury.
Health
Going in to shock can cause renal and liver damage
b/c blood diverted from capillaries to maintain a high
blood presseure.
Varicose veins: from being on feet all day
Elephantitis: lymph fluid doesnt drain out, causes
swelling.
Arteriosclerosis: general hardening of arterial wall.
Atherosclerosis: hardening due to a plaque in the
lumen.


Immunology
Function
Humoral immunity: B cells. Release antibodies in fluids
(guns at rear)
Cell-mediated immunity: T cells. Direct contact (infantry)
Memory (faster response on 2
nd
exposure to antigen)
and specificity for certain antigens.
Tolerance: self from non-self.
Immunoglobulins
Y-shaped. Have heavy and light chains w/ constant and
variable regions. Fab: binds antigens. Fc: binds cell
Help prevent pathogen from binding host cell, promotes
phagocytosis / lysis
IgG: most abundant. Used in 2 response.
IgM: pentamer. On B-cell surface. Used in 1
response.
IgA: dimer. Found in secretions.
IgE: allergic rxns. Makes mast cells & basophils
release granules by binding.
IgD: on B-cell. Not well understood.
Cells
B Lymphocytes
Programmed & mature in bone marrow (1 organ)
Give rise to plasma cells
Only recognize specific antigen once they encounter
it in 2 lymph organs (antigen-dependent
programming).
After encounter, undergo blast transormation &
clonal expansion.
Most clones become effector cells to combat
antigen. Others remain quiescient as memory cells
for faster expansion upon next encounter.
T-Lymphocytes
Precursors programmed in marrow, mature in
thymus (1 organ). Leave as Helper Cells (CD4) and
Cytotoxic Cells (CD8).
Also form effector & memory clones upon encounter
w/ antigen outside thymus.
Natural Killer Cells: Dont need antibodies
Macrophages: presents pieces of pathogen to
lymphocytes so they can recognize it.
MHC1: carried by macrophages on surface. Shown
to CD8 after pathogen destruction.
MHC2: Shown to CD4.
Reticular / Dendritic: holds pieces of pathogens,
stimulates B cells to make antibodies to the pieces.
Holds HIV virus for years, re-relases it once it dies.
Organs
Found in subepithelial CT & lamina propria (provides
blood supply to epithelium)of hollow organs.
Ex: Peyers Patches in GI tract.
Organs encapsulated. Tissues (mase of nodules)
unencapsulated.
Nodules
White germinal center (lymphoblasts) surrounded by
pink mantle (memory cells).
Immunology, Contd
1 Organs: Marrow & Thymus
Thymus
Surrounded by CT capsule. Septae form lobules.
Has epithelial reticular cells (from endoderm), unlike
other lymph organs.
Lobules have cortex (where T-cells programmed) &
medullas (where they enter circulation). Also has
Hassals corpuscles (reticular cells collapse into
areas vacated by T-cells).
Incomplete lobulation: adjacent lobules have their
own cortex, but share a medulla.
Thymus in children is large, easily seen in x-ray.
Shrinks and is replaced by fat w/ age.
Blood-thymus barrier: continuous capillaries +
reticular cells. Prevents antigens from entering and
causing stem cells to prematurely differentiate
(privileged space).
2 Organs
Lymph Nodes
Found in areas where we bend (armpit, groin,
elbows, knees, neck)
Has many nodules. Sinuses filled w/ reticular cells (from
mesoderm) & macrophages
Cortex divided by trabeculae. B-cells divide in white
region of cortex.
T-cells found in paracortical region (btw cortex &
medulla) Enter capillary beds to move into circulation.
80% of lymphocytes in lymph circulation are T-cells. B-
cells stay in node to release antibodies.
Afferent lymph vessel (enter convex side) subcapular
sinus peritrabecular sinus medullary sinus
efferent lymph vessel (leaves node through hilus)





T-cells burrow though high endothelial venules in
paracortex to enter & leave blood circulation. Done
w/ selectin receptors & chemokines for adhesion
(diapedeses)
Antigens & dead cell debris removed in nodes.
Spleen
Inferior tip easily palpated during inflammation.
Removes old RBC (glyococalyx on surface gets worn
off as they age. Spleen targets these).
Can act as a reserve of blood.
Splenic areteries enter though hilus, run through
trabeculae.
Areterioles surrouned by periartiole lymphatic sheath
(made of white pulp & macrophages. Rich in T-cells).
Also surronded by peripheral white pulp (lymphatic
nodules + B-cells).
Empty into red pulp sinuses. Sinuses return blood to
trabecular & splenic venules and veins.
Red pulp = lacks lymphocytes.
Red pulp cords = reticular CT & plasma cells.
Red pulp sinus = endothelial stave cells & basal
lamina.
Tonsils
Partially encapsulated.
Palantine tonsils (back of mouth)
Stratified squamous non-keratinized. Crypts
increase antigen binding surface area. Larger in
children.
Pharyngeal tonsils (nasopharynx)
Ciliated pseudostratified columnar epithelial
Lingual tonsils (back of tongue)
Stratified squamous non-keratinized
All form a protective ring around entrance to
respiratory or digestive system.
Phospholipid Metabolism
Deficient in premature babies. Diabetic mothers at greater
risk (hypoglycemia doesnt induce enzymes for making
surfactant).
Lecithin/sphingo ratio should rise sharply after 32 weeks for
healthy lungs.
Treated w/ Survanta (bovine extract) or Exosurf.
Sphingolipids
Key component of neurons. Palmitoyl-CoA + serine.
Sphingomylein: no glycerol backbone, has long unsat chain
(double bond). Phosphorylated.
Cerbroside: like above, but glycosylated.
Ganglioside (ceramide): have at least 1 NANA (N-acetyl
neuraminic acid). Globoside = no NANA.
Synthesis
Palmitoyl-CoA + serine (some steps) ceramide
Ceramide + p-choline = sphingomylein
Ceramides cerebrosides sulfatides.
Ceramides also make higher gangliosides
No regulation other than availability of substrates.
Degradation
Each terminal sugar is degraded one by one.
Neuraminadase cleaves NANA 1
st
. Ceramide is last to go.
Diseases occur if enzymes in pathway arent present
(intermediates accumulate). See Clinical Correlates!
Leukotrienes & Prostglandins
Synthesis
Phospholipids (Phoshpolipase A2) Arachidonic acid
(cyclooxegenase 1 & 2) Prostglandins
Arachidonic ( 5-lipooxygenase) Leukotrienes
PG and leuko have long, bent unsat chains. PG: pain
response Only in body for ~60 min. Leuko: allergy response
(too much = anaphylactic shock).
Regulation:
A2 inhibited by steroids (cortisol).
COX2 inhibted by NSAIDs (asprin, celebrex). Cause side
effects if they inhibit COX1 (used by body to maintain blood
pressure).
5-lipoox inhibited by zileuton.



Triacylglycerols
Synthesis
1
st
step: Make glycerol phosphate. Made 2 ways:
Use glycerol kinase (in liver, not adipocytes) or glycerol P
dehydro (uses DHAP from glycolysis) Glycerol-P.
Afterwards, becomes triacylglycerol (from 1,2 diacylglycerol).
C1: has saturated FA. C2: unsaturated. C3: either.
Degradation
Gets packaged into VLDL.
All trigly eventually become acetyl-CoA and enters Krebs (if a lot
of -ox occurs, liver turns some to ketone bodies)
Phospholipids
Structure: 2 FA groups attached to one of these phosphatidyl
groups:
P-serine (shortest)
P-ethanolamine
P-choline
P-inositol (cyclic alcohol)
P-glycerol (cardiolipin)
Serine & ethanolamine can interconvert.
Synthesis: 2 key enzymes
Glyc 3P- (several steps)- phosphatidate (CDP diacylgly trans-
ferase / CDP synthase) CDP-diacylglycerol. Makes ALL
phospholipids
(phosphatidate phosphohydrolase) 1,2 diacylglycerol. Makes
ONLY p-choline and p-ethanolamine.
Degradation
4 enzymes: Phospholipases A1, A2, C, and D.
Phospholipases only work on intact phospholipids. After
removing 1
st
FA, lysophospholipase removes 2
nd
FA.
Phospholipase C cleaves p-inositol to make IP3 (affects
cellular Ca receptors).
Lung Surfactant (dipalmitoyllechtin): made 2 ways
Direct acylation: Palmitoyllechtin + Palmityl-CoA = surfactant
[lechtin is same as p-choline]
Exchange Pathway: PalmOL + glycerol p-choline =
surfactant
Prevents alveoli from collapsing.



Genetics
Incomplete penetrance: Person has disease genotype,
but doesnt express it (due to other phenotypic or
genotypic factors). Can still transmit it.
New mutations: Arise due to exogenous factors or abnormal
individual egg or sperm.
Variable expression: disease severity wont be same in all
people due to environment, polymorphisms in regulatory
sequences, or mosaicism.
Locus heterogenity: different genes on different
chromosomes cause same clinical phenotype.
Colorectal cancer due to different MMR genes.
Allelic heterogeneity: different mutations on same gene
(Becker/Duchenne muscular dystrophy)
Uniparental Disomy: both alleles inherited from same parent
Risk increases w/ maternal age.
Gaining 2 alleles from 1 parent + 1 allele from the other
(trisomy) usually fatal before birth.
Mitochondrial Disorders
Heteroplamsy: All mtDNA come from mother, but we
have several populations of mtDNA.
Disease arises if just a few aberrant mitochondria are
passed on to fertilized egg.
Usually affect muscles & nerves.
Multifactorial Traits
Caused by interaction of many genes & environ.
Continuous phenotypes: have Gaussian distribution
(height, BMI, blood pressure)
Often have to pass a threshold before disease trait is
expressed (overweight obese)
Discontinous phenotypes: trait is either present or not
(cleft palate).
Background
From Mendel: genes segereate equally into daughter
cells, & genes from different loci are randomly &
independently assorted.
Inheritance Patterns
Visualized using pedigrees
Single gene (monogenic) inheritance:
Autosomal Dominant
Homozygous Dominants & Heterozygotes display
phenotype. Found in every generation. Both sexes.
Marfans, Huntingtons, Mytonic Dystrophy
Autosomal Recessive
Heterozygotes = carriers. Skips a generation. Both
sexes.
Sickle cell, cystic fibrosis
X-linked Conditions
Males only have one X chromosome, will always
display mutant X. Women will need two mutant Xs
to show disease.
Not transmitted from father to son
X-linked Dominant: Vit-D resistant rickets
X-linked Recessive: G6P-dehydro deficieny
Complications
Lyon Hypothesis/Barr Bodies
Women always inactivate one X chromosome in
each of their cells. Silenced by XIST RNA.
Becomes Barr body (heterochromatin: never
transcribed).
In some cells its maternal X, sometimes paternal.
Women are mosiacs.
Inactivation in women w/ X-linked disorders may
worsen symptoms or have no effect.
Penetrance: Probability that a gene will have a
phenotypic expression.


Antibodies
Synthesis
Gene clusters for each chain found on different
chromosomes
kLight on #2; Light on 22; Heavy on 14.
H chain DNA has V, J, and D sequences (for
varibility). L chain only has V & J. Constant
sequence always at end.
DNA sequences randomly spliced together. Thus, a new
B-cell in marrow will always make Ab that target one
specific antigen, though it hasnt seen any antigens yet.
This process makes us immunocompetent (a healthy
person can respond to nearly any antigen anywhere)
Junctional diversity (how variable sequences are
spliced) is most important factor in conferring diversity.
A small membrane binding DNA sequence at the end
needs to be removed for Ig to be secreted.
Classes
Named for Greek letters (MADGE).
IgM always synthesized 1
st
. Heavy chain DNA sequence
determines Ig class.
Class Switch: APCs & other B-cells stimulate helper T-
cells with their antigen-bound MHCs. Activated T-cells
then release cytokines & interleukins 4 & 5, which start
2
nd
round of DNA rearrangement/splicing in B-cells.
Other Ig classes can now be made.
Ig Superfamily
Made up of other proteins w/ Ig-like domains. Aid in host
defense.
MHC Complex: AKA Human Leukocyte Antigens. 2
classes. Bind & present Ag to T-cells. NOT B-CELLS!
T-Cell Receptors: TcR doesnt bind to antigen directly.
Only binds MHC-Ag complex. CD3 binding helps
activate T-cell.


Terminology
Immunoglobulin (Ig): glycosylated gamma globulin
proteins w/ heavy & light chains.
Antibody (Ab): An Ig that binds a specific antigen (Ag).
Immunogen: an Ag that sets off an immune reaction
upon binding (blasting & cloning)
Structure
Y-shaped. Fab region (binds antigens) on top, Fc
region (binds cell) on bottom.
Joined by hinge region. Cleaved by papain to make 3
fragments. Pepsin seperates Fab from Fc.
2 Light Chains (on outside)
Both chains can either be k or .
Light chains short, dont go past hinge.
Variable region at top of Y (N-terminus), constant
region ends at hinge.
2 Heavy Chains
Variable region at top, constant regions (3 per
chain) go to C-terminus.
Chains joined primarily by disulfide bonds.
Variable regions confer specifity to Fab.
Function
Immune complex: when Ag and Ab bind. Stabilized by
H-bonds & hydrophobic interactions.
Neutralizing: bind pathogens & toxins so they cant bind
to host cell surface.
Promote phagocytosis of antigens by macrophages
(opsonization).
Complement activation: Ab drill holes in an infected
host cell to lyse it. Releases more Ag into circulation to
activate more lymphocytes.
Ab-dependent cellular cytotoxicity: killer T-cells
recognize & bind Ab.
Antigen Presenting Cells (APCs): macrophages &
dendritic cells. Ab enhance their function.



Autonomic Nervous System
Divisions
Parasymp: digest, diuresis, defecation.
Symp: excitement, emergency, exercise.
Receptors
ANS receptors couple to G-proteins, which can set off
cascades in target organs by stimulating secondary
messenger enzymes (adenylate cyclase & phospholipase C).
1: constriction, smooth muscle contraction
intracellular Ca
Sphincters, irises, vascular smooth muscle, skeletal muscle
2: relaxation, releases NE that binds to itself to stop more
NE release (negative feedback on itself).
cAMP
GI tract (relaxes)
1: role specific to location. heart rate, lipolysis,
gluconeogenesis.
cAMP
Heart ( SA/AV node conduction, contractility), kidney, adipose
tissue, salivary gland.
2: dilation (relaxation)
cAMP
vascular smooth muscle
Bladder: relaxes wall & detrusor muscle
Bronchioles: dilation
Liver
Nicotinic: always stimulatory!
Open Na & K channels.
Receptors found on all postganglionic nerves of symp & parasymp
system.
Muscarinic: inhibitory OR stimulatory
intracellular Ca
Sweat glands
Contracts smooth vascular muscles
GI tract (contracts walls, relaxes sphincters, GI secretion)
Relaxes heart rate
Bladder: contracts wall & deterusor skeletal muscle, relaxes
sphincter
Males: erection



Norepi works best w/ receptors, epi works best w/ .
Drugs (w/ site of action)
Symp / parasymp ganglia
Excitatory:
Nicotine
Block AChase:
Physostigmine (Eserine); Neostigmine (Prostigmin)
Block conduction:
Hexamethonium. Wide effects!
Endings of postgang adrenergic nerves
Release norepi
Cocaine; Ephederin; Amphetamine
Stop norepi storage
Resperin
receptors
Stimulatory
Phenylephrine (Neosynephrin); Clonidine; Norepi
Inhibitory
Phenoxybenzamine (Dibenzyline); Phentolamine
(Regitine); Yohimbine
receptors
Stimulatory
Isoproteronol (Isuprel); Epi; Norepi
Inhibitory
Propranolol (Inderal)
Muscarinic receptors
Stimulatory
Ach; Muscarine
Inhibitory
Ach; Atropine; Dramamine; Scopolamine
Nicotinic receptors
Stimulatory
Ach; Nicotine; Carbacol
Inhibitory
Curare; Hexamethonium


Nucleotide Metabolism
Overview
Purine: 6-membered ring + 5-membered ring
A, G, Xanthine (called inosine w/ ribose: short-
lived intermediate)
Pyrimidine: 6-membered ring
T, C, U, Orotic acid
Nucleotide: Base + Sugar + Phosphate (-side = no
phos)
De Novo Purine Synthesis (w/ enzymes)
Ribulose 5P (PPRP synthetase) PPRP (glutamine +
PRPP amidotransferase) PRA (glycine + P-ribosylgly
synth) GAR (Formyltrans + THFolate) FGAR (FGAR
amidotrans) FGAM (AIR synth w/ ATP) AIR (AIR
carboxylase) carb-AIR (ATP + aspartate) SAICAR-
(lyase) AICAR (Formyltrans + THF)- FAICAR
(inosinicase) Inosine (IMP)
Occurs in liver cytosol.
PPRP Synth: rate-limiting step! Enzyme inhibited by
NDP or NTPs
PRPP Amidotransferase: committed step! Dimerized
enzyme = inactive
AIR = closure of 5-membered ring. IMP = closeure of
6-membered ring
Origin of Ring Components
N1: Aspartate
C2, C8: Formyl-THF (from folic acid)
N3, N9: Glutamine
C4, C5, N7: Glycine
C6: CO
2
Inhibitors
Many anticancer & antibacterial drugs interfere w/
cells ability to use THF or glutamine. Stops ring
synthesis.

Azaserine: irreversible glutamine antagonist.
Methotrexate: blocks dihydrofolate reductase, limits THF
synthesis.
Making AMP & GMP
IMP (Adenylsuccinate synthase + GTP + aspartic acid)
Adenylsuccinate (Adenylsuccinase) AMP +
Fumerate
IMP (IMP dehydro + H20 + NAD+) Xanthosine MP
(GMP Synthetase + Glutamine + ATP) GMP
Guanylate & adenylate kinases make GDP & ADP.
Nucleoside diphophate kinase makes GTP & ATP.
Purine Salvage Pathways
Gets purines from diet & normal turnover of old
nucleotides.
Enzymes: Adenine Phosphoribosyl Transferase (makes
AMP) & Hypoxanathine-Guanine PRT (makes IMP &
GMP)
Lesch-Nyhan Syndrome: Inherited X-linked deficiency of
HGPRT. Causes excess uric acid production,
hyperuricemia (gout), involuntary movement, retardation,
self-mutilation.
Purine Degradation
All get broken down to xanthine, which becomes uric acid
Adenine deaminase deficiency: causes buildup of dATP,
which inhibits DNA synthesis.Also causes Severe
Combined Immunodeficincy (SCID: dysfunctional T-cells)
Purine nucleoside phoshporylase deficiency: T-cell
dysfxn. dGTP builds up. PNP turns AMP & GMP to xanth.
Xanathine oxidase inhibition: results from treatment of
gout (Allopurinol removes uric acid crystals, inhibits
oxidase).
Gout treatment: avoid foods w/ a lot of nitrogen (proteins).

N1 C6
N7
N9
Nucleotide Metabolism, Contd
De Novo Pyrimidine Synthesis (w/ enzymes)
2 ATP + CO
2
+ glutamine (Carb-P synth II) Carbomyl
phosphate + Asp (Asp transcarbamoylase) Carb aspartate
(Dihydrooratase) DHO (DHO dehydro) Orotate (O
phosphoribyl trans + PRPP) OMP (Orotidylate decarb)
UMP
CPS II: rate-limiting step
Asp transcarb: commited step.
DHO: ring closed
Pyrimidine ring made before ribose is added.
Orotate (orotic acid): 1
st
base made, short-lived.
Last 2 enzymes are part of same enzymatic complex.
Diseases
Orotic Aciduria: deficincy in OPT & O decarb. Orotic
acid builds in blood. Stops growth, causes
megaloblastic anemia & leukopenia. Mild forms cause
carb-P to build up in liver mitochondria.
Treated w/ oral doses of C or U.
Also, oral UMP converts to UTP, which inhibits
CPS II and stops orotic acid synthesis.
Making CTP & TTP
UTP + H2O + glutamine (CTP synthase) CTP
UDP (Ribonucleotide reductase) dUDP (H
2
O)
dUMP - (Thymidylate synthase) TMP
RR needs thioredoxin & NADPH to make ribose into
deoxyribose (uses H:- radical to remove OH)
TS needs cofactor THF.
Origin of Ring Components
C2 & N3: Carbomyl-P (derived from CO
2
& glutamine)
C4 to N1: Aspartate

Antineoplastic Drugs
5-flourouracil: thyamine analog, inhibits TS.
Aminopterin: like methotrexate. Folate analog, stops
THF synthesis
6-mercaptopurine: inhibits Adenylsuccinate
synthase (IMP to AMP) and IMP dehydro (IMP to
GMP). Also blocks PRPP Amidotransferase (stops
purine synth)
Degradation
Not a lot of disorders seen in pyrimidine breakdown.
Final products are CO
2
+ NH3, which are soluble.
Phosphorylases & kinases act on their respective
bases (Uracil to UMP; Thymine to dTMP;
Deoxycytidine to dCMP has only kinase).
N1
C6
Skin
Info
Largest & heaviest organ
Functions: Protection; Vitamin D manufacture; absorption of
substances; secretion of sweat & sebum; thermoregulation
(metartioles dilate/constrict as needed); sense (touch).
Organization
Epidermis:
Separated from dermis by basal lamina (made up of collagen
IV, lamin, dermatan. Collagen VII fibers used in adhesion).
Many strata. Keratinoytes fill up w/ keratin as they move
higher.
Germanitivum or basale: bottom layer
Spinosum: next layer up. Connected by desmosomes
(spines). Has Langerhans cells (antigen-presenting).
Spinosum & germanitivum = stratum malphigi
Granulosom: large, irregular shaped. 3-5 layers thick. Seals
out water.
Lucidum: thin, translucent layer. Cells have immature keratin
called eledin.
Corneum: highest layer. Has lots of keratin.

Dermis
Dermal papilla ("hills"): extend up into epidermis, form
epidermal ridges ("valleys"). Dermis has blood vessels while
epidermis has none.
Upper dermis (papillary): loose CT, collagen III. Lower dermis
(reticular): dense CT, collagen I.

Pigmentation
Melanin: UV protection. Made from tyrosine by enzyme
tyrosinase. W/out it, makes skin and irises very fair.
Melanocytes: found in stratum germanitivum. Use cytocrine
to secrete melanin to other cells.



Caroten: makes skin slightly yellow.
Oxyhemoglobin (blood): makes skin slightly red.
Nails
Nail plate lies on top of nail bed. Grows from
matrix
Eponchyum: region of nail by cuticle.
Hyonchyum: region farthest from cuticle
(fingertip).
Hair
Starting from inside: medulla, cortex, and cuticle.
Surrounded by internal & external root sheath.
Phases: growing (anagen), regression
(catagen), resting (telogen).
Glands
Sebeacous glands: located by hair follicles
Sweat glands: Eccrine clear, watery, odorless.
Apocrine pigmented, thicker, odor.
Diseases
Burn blisters: occur when epidermis is separated
from dermis.
Pemphigus: autoimmune disorder. IgG attacks
desmosomes in epidermis.
Pemphigoid: autoimmune disorder. Occurs in
basement membrane (dermis). Sub-epidermal.
Psoriasis: Patches of hyper-keratinized skin
(keratinocytes migrate to surface too fast)
Ichtyosis: skin produces too much keratin,
appears rough & scale-like.

Spinal Cord and Nerves
Disorders
Spina Bifada
Occulta: Vertebral arch not fully closed, maeked
by tuft of hair. Usually 1 or 2 lumbar vertebrae.
No symptoms. ~10% of popularion.
Meningocele: sunarachnoid space protrudes
out of spinal column, but spinal cord remains
inside. ~1/1000 births.
Meningomyelocele: spinal cord displaced.
Myelocele (aka myeloshisis or rachischisis):
spinal cord split open, flat like a book. No neural
tube formed. Fatal.
Cystic forms associated w/ elevated levels of
fetoprotein. Intake of folic acid must be
adequate during 3
rd
& 4
th
week to prevent this
Slipped Discs (hernias): if posterior longitudinal
ligament of vertebrae isnt strong, disc can slip out,
pinch spinal cord.
Whiplash: atlanto-axial joint (1
st
& 2
nd
vertebrae) in
neck is affected, compresses spinal cord.
Axis has protrusion called dens. fits tightly into
the atlas above it.
Divisions
Central Nervous System
Brain & Spinal Cord
Peripheral Nerves
Cranial nerves (12 pairs)
Spinal nerves (31 pairs)
Motor & Sensory Neurons
Afferent: Sensory
Pseudounipolar (cell body in ganglion)
General sense: widespread phenomenon (i.e. touch)
vs. special sense (vision, taste)
Somatic: body wall & limbs
Visceral: internal organs
GSA & GVA enter dorsal spinal cord.
Efferent: Motor neurons
Control contraction and secretion
Multipolar
Somatic: skeletal muscle, spindles.
Visceral: smooth & cardiac muscle, glands (typically
via 2 neuron pathways)
GSE & GVE enter ventral spinal cord.
Development
Innervate myotomes, which split into epimere &
hypomere.
1 ramus (branch after spinal nerve): Dorsal innervates
epi, ventral innervates hypo.
Muscles migrate to different parts of body during
development, but always keep their 1
st
nerves.
The Back
Bones
33 vertebrae (7 cervical, 12 thoracic, 5 lumbar, 5 fused
sacral, 3-5 fused coccyx)
1 curvature: thoracic & sacral. Convex posterior.
2 curvature: develop after birth. Cervical & lumbar. Concave
posterior.
Contains internal vertebral venous plexus: main pathway for
tumors to metastisize upper regions of body.
Muscles
Superficial & intermediate muscles
Innervated by ventral 1 rami. Derived from hypomere . Formed
laterally, migrated to back.
Deep (intrinsic) muscles
Innervated by dorsal 1 rami. Derived from epimere.
Nerves
Plexi: ventral rami from different vertebrae intermingle. Each
ramus nerve can contribute to many different target muscles.
Ventral rami of thoracic body wall distribute as simple segments:
discrete, dont form plexi.
Conus medullarus: end of spinal cord. Usually btw T12 & L3.
Gives rise to caudae equina, long dorsal & ventral roots of
spinal nerves that strech out from conus before exiting spine.
Slipped disks: N + 1 Rule
Cervical region: nerves exit above their named vertebrae
(8 spinal nerves: last one exits above T1)
Lumbar reagion: caudal nerves their named vertebrae at
same level as disc, but exit spine at sharp angles. Discs
will miss their own nerves, but will pinch the nerve from
above.



Dermatome: nerves (plexi or simple segments)
from a spinal segment innervate an area of skin
in a predictable way
NOT the same as cutaneous distribution of
peripheral nerves (these often recive
contribution from several spinal segments
due to plexi).
T4: skin around nipples
T6: skin around thumb/posterior arm
T10: skin around navel
L4: skin around knee/shin/thigh
More Development Notes
Ribs and vertebrae derived from sclerotomes.
Each vertebrae formed by two neighboring
sclerotomes.
Notochord forms nucleus pulposous in discs.
Pulp acts like a spring, but can't be compressed
or stretched too much or disc could pop out.
Annulous fibrous (from sclerotome)surrounds
nucleus pulposous.
Vertebrae and disc it sits on share same
number.
Amino Acid Metabolism
Background
A.A. metab consumes a lot of NRG. ~30% of resting NRG use.
A.A. dont get stored. Catabolized immediately to get rid of
ammonia. Need to be continuosly replenished from diet.
Ammonia toxic to brain: crosses blood-brain barrier. Combines w/
-keto to make glutamate. -keto needed for Krebs, brain cant
undergo ox phos. No NRG to function! Can cause ataxia, coma.
Glutamate is also a neuro trans, used to make GABA. If too
much ammonia comes in, it takes away available glutamate
to make glutamine.
Retain nitrogen during growth, pregnancy, or injury recovery.
Some proteins have fast turnover (Hb); others stay in body for
years (collagen).
A.A. Properties
Essential a.a. WTF HaVe MILK. Cant be made in body b/c we
cant make their C skeletons.
Lys and Thr cant be made, even if we do eat their C
skeleton.
Arg is conditionally essential (we make it, but its usually not
enough: gets used to synthesize other things).
Tyr: can be made from Phe.
Proline: imino acid instead of an amino acid.
Digestion
Stomach acid denatures proteins
Pepsin (in stomach): endopeptidase. Releases a.a. from interior
of chain.
Secretin & CCK (released by intestine) stimulate pancreas to
secrete basic pancreatic juice
Juice has trypsin, chymotrypsin, elastase. Carboxypeptidase is
an exopeptidase. Enteropeptidase activates typsinogen.
All made as inactive zymogens so they wont digest GI tract
Oligopeptidases: work on chains smaller than 25 a.a.
A.A. taken tnto intestines by Na-cotransporters. Enters blood via
passive facilitated transport.


Diseases
Cystic fibrosis: not enough pancreatic enzymes. Foul-
smelling oily stools, poor growth.
Cystinuria: cant transport a.a. Genetic. Defective renal re-
absorption of arg, lys, orn,cyst, excreted in urine. Also
cause kidney stones
Hartnup: similar to pellagra (dermititis, dementia, diarrehia
& death), but w/ only mild forms of the 1
st
two symptoms.
Cant absorb a.a. Treated w/ niacin.
A.A. Fates
57% oxidized in liver.
23% pass through liver intact to muscle. Mainly
hydrophobic (branched) a.a Leu, Ile, Val.
14% make liver proteins.
6% make blood proteins.
Removing Ammonia
Ammonia always removed (80% as urea, 20% directly in
urine), C skeleton enters Krebs.
Oxidative deamination: Glutamate Dehydro rxn
(reversible): loses NH4 to make -keto. Only a.a. that can
do this!
Glutamate also accepts NH4 to become glutamine. Carries
ammonia through blood to liver. Rxn done by glutamine
synthetase, needs NRG.
Glutaminase Rxn (hydrolysis): Turns glutamine to
glutamate, releases ammonia.
Glutamine cycle scavenges any free ammonia missed by
urea cycle. Also reatains bicarbonate (basic) during blood
acidosis rather than excreting the bicarbonate in urine.
Alanines C skeleton=pyruvate! Aspertatess C
skeleton=oxaloacetate! Both formed after transamination
w/ glutamate (forms -keto!)
Asp can form asparginine w/ addition of another
nitrogen group. Releases it w/ asparginase.


Amino Acid Metabolism, Contd
Remember: transamination from one a.a. to another is
a ping-ping mechanism! Asp & alanine
aminotransferases use cofactor Vit-B6 (pyridoxal
phosphate).
B6 vital for all a.a. metabolism!
Urea Cycle
NH4 + Bicarbonate + 2 ATP (CPS1) CP + Ornithine
(OTC) Citrulline + Asp + ATP (Arg-Succinate
Synth) Argsuccinate (AS Lyase) Arg + Fumerate
(Arginase) Ornithine + Urea
NAG is important allosteric activator of CPS1.
Made from Glutamate + Acetyl-CoA.
CPS1: rate limiting step.
CPS1 & OTC in liver mitochondria! Citrulline diffuses
out to cytosol, ornithine diffuses back in!
Glutamate supples all the ammonia.
Fumerate later becomes Asp.
Arg also makes NO (vasodilator & anti-inflammatory).
Diseases
Type I Hyperammonemia: deficiency in CPS1. Rare.
Type II Hyperammonemia: deficincy in OTC. X-linked.
Citrullinuria: deficiency in ASS.
Arginosuccinic Acidemia: deficiency in ASL.
Hyperargininemia: deficiency in arginase.
NAGS deficiency VERY serious: large amounts of NH4
in body.
Hyperammonemia can also occur during cirrohsis.
Categories
Strictly ketogenic: Leu & Lys (LK)
Ketogenic & glucogenic: Ile, Phe, Thr, Trp, Tyr (IFTWY)
Strictly glucogenic: all others.


Fates of C Skeletons
Oxaloacetate: made from Asp & Asn
-keto: from Glu, Gln, Pro, Arg, His
Histidinemia (usually beningn) & folic acid deficiency
Pyruvate: from Ala, Ser, Gly, Cys, Thr, Trp
Ser & Thr dehydratases use B6 as cofactor.
Homocysteinuria: no cystathionine synth. Can also be
caused by lack of folate or B12.
Amino oxidase turns gly to oxalate. Deficiency =
hyperoxaluria: oxalate build up & kidney stones.
Fumerate: from Phe & Tyr
Phe hydroxylase needs biopterin as cofactor
Alkaptonuria: no homogentisic acid oxidase (cant break
down Tyr). Causes arthritis & black urine.
PKU: most common a.a. metabolic disorder! Deficiency
of Phe hydroxylase. Phe buildup, causes hypo-
pigmentation & neural problems. Needs biopterin.
Albinism: no tyrosinase: cant make melanin.
Remember! Phe (essential) makes Tyr!
Succinyl-CoA: from Met, Thr, Ile, Val.
Met makes propionyl CoA (succ intermediate), needs
biotin, B12, B6
Methyl malonyl CoA mutase deficiency: causes
ketoacidosis, short stature, retardation. Needs B12.
Acetyl-CoA: from Leu, Ile, Lys, Try
Pellagra: Deficiency in Trp. Treated w/ niacin.
Branched Chain: Leu, Val, Ile
Isovaleria: build up of isovaleryl-CoA, (faulty Leu
degradation)
Maple Syrup Urine Disease: deficiency in branch chain
-keto acid dehydro. Ketoacid buildup. Can be lethal if
not detected early. Needs B1 (thiamine).