DNA and RNA Structures

DNA AS GENETIC
MATERIAL
01/11/17
It is well known fact that transmission of traits takes place

from one generation to other. The offsprings are similar to
both the parents in some traits
Gregor Johann Mendel (1866 ) on the basis of his
hybridization experiments on Sweet pea gave the idea that
transmission of traits over generations take place through
Factor or Determiner or Gene which carries information for
expression of trait or phenotype.
Genes are present on the chromosomes which are distributed
equally into the two daughter cells during cell division. The
biochemical studies reveal that chromosomes are composed
of proteins (60% ) and DNA (40% )
01/11/17
Genetic
capable of
material
must
be
Replication ( Make its copy )

Storage of information for expression of
trait
Control expression of traits
Change in controlled way (undergo
mutation)
Must be stable
01/11/17
On the basis of certain experiments conducted from time to

time ,it was ultimately demonstrated that DNA
carries
genetic information and not the proteins

There are some direct evidences
and some indirect
evidences which prove DNA as Genetic Material
Directevidencescomefrom:
Frederick Griffiths (1928) experiment on Bacterial
Transformation
Oswald Avery, Colin Macleod and Maclyn McCartys
(1944 ) experiment on Transformation
Alfred D. Hershey and Martha Chase (1952 )
experiment on T- Even (2,4 ) Bacteriophage
01/11/17
Frederick Griffith (1928) Studied Diplococcus

pneumoniae, having Two strains
SIII strain was virulent, possessed a lipopolysaccharide
capsule
and
could
kill
mice
by
causing
disease
Pneumonia and made round colonies on a culture plate

RII strain was avirulent and lacked a Lipopolysaccharide
(LPS) capsule, growing in roughshaped colonies on a
culture plate
01/11/17
01/11/17
STEPS IN THE
EXPERIMENT
1 LIVE
SIII
2 LIVE
RII
3 HK S
III
4 H K S III
&
Strains of Diplococcus pneumoniae injected LIVE RII
to mice
01/11/17
Griffiths
Experiment
RII SIII
transformatio
n takes place
in step 4
give clue for
DNA as
genetic
material
4
01/11/17
STEPS AND RESULTS OF GRIFFITHS

EXPERIMENT
SN
RESULT
I
II
III
Mouse injected with SIII strain

Mouse died
Mouse injected with RII strain
Mouse survived
Mouse injected with Heat Killed SIII Mouse survived
strain
IV
Mouse injected with Heat Killed SIII Mouse died & from
& living R II strain
its blood live SIII
VI
01/11/17
STEPS
Mouse injected with Heat Killed

SIII+ living RII strain + DNase
enzyme
Mouse injected with Heat Killed
SIII+ living RII strain + Protease
enzyme
strain bacteria
recovered
Mouse survived
Mouse died of
Pneumonia
9
GRIFFITHS
CONCLUSION
On the basis of Result of step IV Griffith concluded that there

was transformation of Avirulent RII type to Virulent SIII type
by picking up the genetic material encoding the LPS capsule
from the Heat Killed S III .
This bacterial transformation clearly shows the role of DNA as
Genetic Material and is further confirmed by results of step V
which shows no transformation as DNase digests DNA and
step VI again shows transformation as protease only digests
protein
01/11/17
10
Avery,
MacLeod,
&
McCarty,
1944
repeated
Griffiths
experiment of transformation using purified cell extracts and

concluded
Removal of all protein from the transforming material did not
destroy its ability to transform R strain cells
DNA-digesting enzymes destroyed all transforming ability
The transforming material is DNA
01/11/17
11
AVERY,MA
CLEOD
AND
MCCARTY
(1944)
01/11/17
12
Avery, MacLeod AND McCarty

EXPERIMENT
Is based on transformation
Cell free extract of SIII strain Bacterium was subjected to

DNase,RNase and Protease
Each treated extract was mixed with RII and mixture

injected to mouse to see transformation.
In case of Protease and RNase transformation was
recorded
In case of DNase no transformation was recorded
01/11/17
13
Avery et al (1944) revealed the chemical nature of

the transforming substance to be DNA. With the help
of experiment they showed that DNA isolated from
SIII strain Bacteria could confer the pathogenic
properties to R II strain Bacteria .
Two conclusion were derived
1.Active
factor
is
DNA
which
can
cause
transformation
2.SIII strain contains the Active factor
01/11/17
14
THE MODERN FRAME WORK OF

MOLECULAR BIOLOGY
Transcription
Processing
Translation
DNApreRNARNA
Protein
Replication
&Repair
DNA
CentralDogma
01/11/17
Folding
Assembly
Processing
15
DNA Structure,
Functions and
Properties
BiochemistryforMedics
16
DNA
DNA - a polymer of deoxyribo
nucleotides
found in chromosomes, mitochondria and
chloroplasts
carries the genetic information
17
Components of a nucleotide
Base
Sugar
Phosphate
18
Base
Phosphate
Sugar
X=H: DNA
X=OH: RNA
Nucleoside
Nucleotide
19
Basic structure of
pyrimidine and purine
20
Pyrimidines
21
Purines
22
Nomenclature of Nucleic Acid Components

Base
Purines
Adenine
Guanine
Pyrimidines
Cytosine
Thymine
Uracil
Nucleoside
Nucleotide
Nucleic
acid
Adenosine
Deoxyadenosine
Guanosine
Deoxy guanosine
Adenylate
Deoxyadenylate
Guanylate
Deoxyguanylate
RNA
DNA
RNA
DNA
Cytidine
Deoxycytidine
Thymidine
(deoxythymidine)
Uridine
Cytidylate
Deoxycytidylate
Thymidylate
(deoxythymidylate)
Uridylate
RNA
DNA
DNA
RNA
23
Each strand
consists of:
1) A Sugar Phosphate Backbone
Each strand
consists of:
1) A Sugar Phosphate Backbone

2) Four Base Chemicals
(Attached in pairs)
Adenine pairs with Thymine

Guanine pairs with Cytosine
Structure
BasicstructureofDNAisasugarphosphatebackbonewith4
variablenitrogenousbases.Thisstructureiscalleda
nucleotide.
P
Phosphate
molecule:
HYDROPHILIC
sugar
5carbon
sugar:
DEOXYRIB
OSE
BACKBONE
Nitrogen
base
Nitrogen
base:
HYDROPH
BASE
OBIC
The primary structure of

DNA is the sequence
5 end
3
Phosphodiester
linkage
3 end
28
Traditionally, a DNA sequence is

drawn from 5 to 3 end.
A shorthand notation for this
sequence is ACGTA
29
The secondary structure of

DNA is the double helix
30
The secondary structure of

DNA
Two anti-parallel polynucleotide
chains wound around the same axis.
Sugar-phosphate chains wrap
around the periphery.
Bases (A, T, C and G) occupy the
core, forming complementary A T
and G C Watson-Crick base pairs.
31
The DNA double helix is held together

mainly by- Hydrogen bonds
hydrogen bonding;
base stacking
32
Hydrogen bond
a chemical bond in which a hydrogen atom of
one molecule is attracted to an electronegative
atom, especially a nitrogen, oxygen, or fluorine
atom, usually of another molecule.
33
Two hydrogen bonds between A:T pairs

Three hydrogen bonds between C: G paired
34
Base Stacking
The bases in DNA are
planar and have a
tendency to "stack".
Major stacking forces:
hydrophobic interaction
van der Waals forces.
35
Structural forms of DNA

Property
A-DNA
B-DNA
Z-DNA
Helix
Handedness
Right
Right
Left
Base Pairs per

turn
11
10.4
12
Rise per base

pair along axis
0.23nm
0.34nm
0.38nm
Pitch
2.46nm
3.40nm
4.56nm
Diameter
2.55nm
2.37nm
1.84nm
Conformation
of Glycosidic
bond
anti
anti
Alternating
anti and syn
Major Groove
Present
Present
Absent
Minor Groove
Present
Present
Deep cleft
36
Normally hydrated DNA: B-form DNA

Helical sense: right handed
Base pairs: almost
perpendicular to the helix
axis; 3.4 apart
One turn of the helix: 36
; ~10.4 base pairs
Minor groove: 12 across
Major groove: 22 across
37
In eukaryotic cells,
DNA is folded into chromatin
38
DNA Tertiary Structure

DNA DOUBLE HELICAL STRUCTURE COILS ROUND
HISTONES.
DNA BOUND TO HISTONES FORMS
NUCLEOSOMES (10nm FIBRES)
NUCLEOSOMES CONTAIN 146 NUCLEOTIDES
39
Nucleosomes
any of the repeating globular subunits of
chromatin that consist of a complex
of DNA and histone
40
Structure of nucleosome core
41
Compaction of DNA in a eukaryotic chromosome
42
DNA melting and annealing
43
Reversible denaturation and annealing of DNA
44
Melting point (tm) of DNA

The temperature at the mid-point of the transition
45
The tm of DNA depends on:
Content of GC base pairs

size of DNA
pH
ionic strength
46
BiochemistryForMedics
47
RNA
is a polymer of
ribonucleotides linked
together by 3-5
phosphodiester
linkage
48
RNA V/S DNA
49
S.No. RNA
DNA
1)
Single stranded mainly

except when self
complementary
sequences are there it
forms a double stranded
structure (Hair pin
structure)
Double stranded (Except

for certain viral DNA s
which are single
stranded)
2)
Ribose is the main sugar
The sugar moiety is

deoxy ribose
3)
Pyrimidine components
differ. Thymine is never
found(Except
tRNA)
Thymine is always there

but uracil is never found
4)
Being single stranded

structure- It does not
follow Chargaffs rule
It does follow Chargaff's

rule. The total purine
content in a double
BiochemistryForMedics stranded DNA is always
50
S.No. RNA
DNA
5)
RNA can be easily

destroyed by alkalies to
cyclic diesters of mono
nucleotides.
DNA resists alkali action

due to the absence of
OH group at 2 position
6)
RNA is a relatively a labile

molecule, undergoes easy
and spontaneous
degradation
DNA is a stable
molecule. The
spontaneous
degradation is very 2
slow. The genetic
information can be
stored for years
together without any
change.
7)
Mainly cytoplasmic, but

also present in nucleus
(primary transcript and
small nuclear RNA)
Mainly found in nucleus,

extra nuclear DNA is
found in mitochondria,
and plasmids etc
8)
The base content varies
Millions of base pairs are
51
S.No. RNA
DNA
9)
There are various types of

RNA mRNA, r RNA, t
RNA, Sn RNA, Si RNA, mi
RNA and hn RNA. These
RNAs perform different
and specific functions.
DNA is always of one

type and performs the
function of storage and
transfer of genetic
information.
10)
No variable physiological
forms of RNA are found.
The different types of RNA
do not change their forms
There are variable forms

of DNA (A to E and Z)
11)
RNA is synthesized from

DNA, it can not form
DNA(except by the action
of reverse transcriptase).
It can not duplicate
(except in certain viruses
where it is a genomic
material )
DNA can form DNA by

replication, it can also
form RNA by
transcription.
52
In all prokaryotic and eukaryotic organisms, three

main classes of RNA molecules exist1)Messenger RNA(m RNA)
2)Transfer RNA (t RNA)
3)Ribosomal RNA (r RNA)
The other are
osmall nuclear RNA (SnRNA),
omicro RNA(mi RNA) and
osmall interfering RNA(Si RNA) and
oheterogeneous nuclear RNA (hnRNA).
53
Comprises
only 5% of the RNA in the cell

Most heterogeneous in size and base
sequence
All members of the class function as
messengers carrying the information in a
gene to the protein synthesizing machinery
54
The
5 terminal end is capped by 7- methyl

guanosine triphosphate cap.
The cap is involved in the recognition of mRNA
by the translating machinery
It stabilizes m RNA by protecting it from 5
exonuclease
55
The 3end of most m-RNAs have a polymer of

Adenylate residues( 20-250)
The tail prevents the attack by 3 exonucleases
Histones and interferons do not contain poly A tails
On both 5 and 3 end there are non coding
sequences which are not translated (NCS)
The intervening region between non coding
sequences present between 5 and 3 end is called
coding region. This region encodes for the synthesis
of a protein.
56
Structural
Characteristics of
m-RNA
5 cap and 3 tail impart stability to m RNA by

protecting from specific exo nucleases.
57
The
m- RNA molecules are formed with the help of

DNA template during the process of transcription.
The sequence of nucleotides in m RNA is
complementary to the sequence of nucleotides on
template DNA.
The sequence carried on m -RNA is read in the form of
codons.
A codon is made up of 3 nucleotides
The m-RNA is formed after processing of
heterogeneous nuclear RNA
58
In mammalian nuclei , hnRNA is the immediate

product of gene transcription
The nuclear product is heterogeneous in size
(Variable) and is very large.
Molecular weight may be more than 107, while the
molecular weight of m RNA is less than 2x 106
75 % of hnRNA is degraded in the nucleus, only
25% is processed to mature
m RNA
59
Heterogeneous nuclear
RNA
(hnRNA)
Mature m RNA is formed from primary transcript

by capping, tailing, splicing and base
modification.
60
Transfer
RNA are the smallest of three major

species of RNA molecules
They have 74-95 nucleotide residues
They are synthesized by the nuclear
processing of a precursor molecule
They transfer the amino acids from cytoplasm
to the protein synthesizing machinery, hence
the name t RNA.
They are easily soluble , hence called Soluble
RNA or s RNA
They are also called Adapter molecules, since
they act as adapters for the translation of the
sequence of nucleotides of the m RNA in to
specific amino acids
There are at least 20 species of t RNA one
corresponding to each of the 20 amino acids
required for protein synthesis.
61
1) Primary structure- The nucleotide sequence of all

the t RNA molecules allows extensive intrastand
complimentarity that generates a secondary structure.
2) Secondary structure- Each single t- RNA shows
extensive internal base pairing and acquires a clover
leaf like structure. The structure is stabilized by
hydrogen bonding between the bases and is a
consistent feature.
62
Secondary structure (Clover leaf structure)

All t-RNA contain 5 main arms or loops which are as
followsa)Acceptor arm
b)Anticodon arm
c)D HU arm
d)T C arm
e)Extra arm
63
a)
Acceptor arm
The acceptor arm is at 3 end
It has 7 base pairs
The end sequence is unpaired Cytosine, CytosineAdenine at the 3 end
The 3 OH group terminal of Adenine binds with
carboxyl group of amino acids
The t RNA bound with amino acid is called Amino
acyl t RNA
CCA attachment is done post transcriptionally
64
Secondary structure of
t- RNA
The carboxyl group of amino acid is attached to 3OH group of Adenine

nucleotide of the acceptor arm. The anticodon arm base pairs with the codon
present on the m- RNA
65
b) Anticodon arm
Lies at the opposite end of acceptor arm
5 base pairs long
Recognizes the triplet codon present in the m RNA
Base sequence of anticodon arm is complementary to
the base sequence of m RNA codon.
Due to complimentarity it can bind specifically with m
RNA by hydrogen bonds.
66
c) DHU arm
It has 3-4 base pairs
Serves as the recognition site for the enzyme (amino
acyl t RNA synthetase) that adds the amino acid to the
acceptor arm.
d) TC arm
This arm is opposite to DHU arm
Since it contains pseudo uridine that is why it is so
named
It is involved in the binding of t RNA to the ribosomes
67
e) Extra arm or Variable arm

About 75 % of t RNA molecules possess a short
extra arm
If about 3-5 base pairs are present the t-RNA is
said to be belonging to class 1. Majority t -RNA
belong to class 1.
The t RNA belonging to class 2 have long extra
arm, 13-21 base pairs in length.
68
The L shaped tertiary structure

is formed by further folding of the
clover leaf due to hydrogen bonds
between T and D arms.
The base paired double helical
stems get arranged in to two
double helical columns, continuous
and perpendicular to one another.
69
The mammalian ribosome contains two major

nucleoprotein subunitsa larger one with a molecular
weight of 2.8 x 106 (60S) and a smaller subunit with a
molecular weight of 1.4 x 106 (40S).
The 60S subunit contains a 5S ribosomal RNA (rRNA),
a 5.8S rRNA, and a 28S rRNA; there are also probably
more than 50 specific polypeptides.
The 40S subunit is smaller and contains a single 18S
rRNA and approximately 30 distinct polypeptide chains.
All of the ribosomal RNA molecules except the 5S
rRNA are processed from a single 45S precursor RNA
molecule in the nucleolus .
5S rRNA is independently transcribed.
70
Ribosomal RNA
(rRNA)
71
The
functions of the ribosomal RNA molecules in the

ribosomal particle are not fully understood, but they
are necessary for ribosomal assembly and seem to
play key roles in the binding of mRNA to ribosomes and
its translation
Recent studies suggest that an rRNA component
performs the peptidyl transferase activity and thus is
an enzyme (a ribozyme).
72
Most
of these molecules are complexed with

proteins to form ribonucleoproteins and are
distributed in the nucleus, in the cytoplasm, or in
both.
They range in size from 20 to 300 nucleotides and
are present in 100,0001,000,000 copies per cell.
73
snRNAs,
a subset of the small RNAs,

are significantly involved in mRNA
processing and gene regulation
Of the several snRNAs, U1, U2, U4,
U5, and U6 are involved in intron
removal and the processing of hnRNA
into mRNA
The U7 snRNA is involved in
production of the correct 3' ends of
histone mRNAwhich lacks a poly(A)
74
Small Nuclear RNAs

(snRNAs).
Sn RNA s are involved in the process of splicing (intron removal) of

primary transcript to form mature m RNA. The Sn RNA s form complexes
with proteins to form Ribonucleoprotein particles called snRNPs
75
These
two classes of RNAs represent a subset of small

RNAs; both play important roles in gene regulation.
miRNAs and siRNAs cause inhibition of gene
expression by decreasing specific protein production albeit
apparently via distinct mechanisms
76
miRNAs
are typically 2125 nucleotides in length and

are generated by nucleolytic processing of the products
of distinct genes/transcription units
The small processed mature miRNAs typically
hybridize, via the formation of imperfect RNARNA duplexes within the 3'-untranslated regions
of specific target mRNAs, leading via unknown
mechanisms to translation arrest.
77
Micro RNAs (miRNAs)
microRNAs, short non-coding RNAs present in all living organisms,

have been shown to regulate the expression of at least half of all
human genes. These single-stranded RNAs exert their regulatory
action by binding messenger RNAs and preventing their translation
into proteins.
78
siRNAs are derived by the specific nucleolytic

cleavage of larger, double-stranded RNAs to again form
small 2125 nucleotide-long products.
These short siRNAs usually form perfect RNA-RNA
hybrids with their distinct targets potentially anywhere
within the length of the mRNA where the
complementary sequence exists.
Formation of such RNA-RNA duplexes between siRNA
and mRNA results in reduced specific protein
production because the siRNA-mRNA complexes are
degraded by dedicated nucleolytic machinery;
some or all of this mRNA degradation occurs in
specific organelles termed P bodies.
79
Small Interfering RNAs

(siRNAs)
Small interfering RNA (siRNA) are 20-25 nucleotide-long double-stranded

RNA molecules that have a variety of roles in the cell. They are involved in
the RNA interference (RNAi) pathway, where it interferes with the
expression of a specific gene by hybridizing to its corresponding RNA
sequence in the target mRNA. This then activates the degrading
mRNA.Once the target mRNA is degraded, the mRNA cannot be
translated into protein.
80
Both
miRNAs and siRNAs represent exciting new

potential targets for therapeutic drug
development in humans.
In addition, siRNAs are frequently used to decrease or
"knock-down" specific protein levels in experimental
procedures in the laboratory, an extremely useful and
powerful alternative to gene-knockout technology.
81

DNA and RNA Structures

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DNA AS GENETIC

It is well known fact that transmission of traits takes place

Replication ( Make its copy )

Change in controlled way (undergo

On the basis of certain experiments conducted from time to

genetic information and not the proteins

and some indirect

evidences which prove DNA as Genetic Material

Frederick Griffith (1928) Studied Diplococcus

Pneumonia and made round colonies on a culture plate

STEPS AND RESULTS OF GRIFFITHS

Mouse injected with SIII strain

Mouse injected with Heat Killed

On the basis of Result of step IV Griffith concluded that there

experiment of transformation using purified cell extracts and

Avery, MacLeod AND McCarty

Cell free extract of SIII strain Bacterium was subjected to

Each treated extract was mixed with RII and mixture

Avery et al (1944) revealed the chemical nature of

THE MODERN FRAME WORK OF

Nomenclature of Nucleic Acid Components

1) A Sugar Phosphate Backbone

1) A Sugar Phosphate Backbone

Adenine pairs with Thymine

The primary structure of

Traditionally, a DNA sequence is

The secondary structure of

The secondary structure of

The DNA double helix is held together

Two hydrogen bonds between A:T pairs

Structural forms of DNA

Base Pairs per

Rise per base

Normally hydrated DNA: B-form DNA

DNA Tertiary Structure

Structure of nucleosome core

Compaction of DNA in a eukaryotic chromosome

DNA melting and annealing

Reversible denaturation and annealing of DNA

Melting point (tm) of DNA

The tm of DNA depends on:

Content of GC base pairs

RNA V/S DNA

Single stranded mainly

Double stranded (Except

Ribose is the main sugar

The sugar moiety is

Thymine is always there

Being single stranded

It does follow Chargaff's

RNA can be easily

DNA resists alkali action

RNA is a relatively a labile

Mainly cytoplasmic, but

Mainly found in nucleus,

There are various types of

DNA is always of one

There are variable forms

RNA is synthesized from

DNA can form DNA by

In all prokaryotic and eukaryotic organisms, three

only 5% of the RNA in the cell

5 terminal end is capped by 7- methyl

The 3end of most m-RNAs have a polymer of