Chapter 4

The Three-Dimensional
Structure of Proteins

Part 1
 Proteins: Structure, Function, Folding

Learning goals: to Know:

Structure and properties of the peptide bond
Structural hierarchy in proteins
Structure and function of fibrous proteins
Structure analysis of globular proteins
Protein folding and denaturation
 Structure of Proteins

Unlike most organic polymers, protein

molecules adopt a specific three-
dimensional conformation.
This structure is able to fulfill a specific
biological function
This structure is called the native fold
The native fold has a large number of
favorable interactions within the protein
There is a cost in conformational entropy of
folding the protein into one specific native
fold
 Favorable Interactions in Proteins

Hydrophobic effect
Release of water molecules from the structured solvation layer
around the molecule as protein folds increases the net entropy
Hydrogen bonds
Interaction of N-H and C=O of the peptide bond leads to local
regular structures such as -helices and -sheets
London dispersion
Medium-range weak attraction between all atoms contributes
significantly to the stability in the interior of the protein
Electrostatic interactions
Long-range strong interactions between permanently charged
groups
Salt-bridges, esp. buried in the hydrophobic environment strongly
stabilize the protein
Folding Final Structure: Chymotrypsin and
Glycine

75 Daltons

21,000 Daltons, 3 polypeptides

 Protein Conformation
Stabilized by weak bonds:
G separating folded and unfolded is small
Bond: H bonds, hydrophobic interatction, ionic
interaction and S-S-.
Proteins possess a solvation layer
The extent of which depends on surface amino
acid R groups

Overall structural patterns:

Hydrophobic areas buried in protein interior
Number of H-bonds is maximized
The Peptide Bond
Only non-planar Bonds Rotate

Each -Carbon has a and

Getting the Angles
Many Angles are Prohibited due to Steric Overlap
Ramachandran Plot
Pauling and Corey and the Alpha Helix
 How To Determine Protein Structure
The Classic Method X-ray Crystallography

Methods to form Crystals take Proteins to their Solubility Minimum

Proteins Crystals in Electron Microscopy
X-ray Diffraction
X-ray Diffraction Pattern of Myoglobin and DNA

Fourier Transform to convert X-ray pattern to Electron Density Map

X ray Diffraction Patterns from Different Proteins
Fitting Electron Density Map to Structure
Fitting Electron Density Map to Primary Structure
X-ray Crystallography at Fine Resolution
Proton Nuclear Magnetic Resonance of a Protein
2 Dimensional NMR
NMR Structure of Myoglobin

NMR is limited to
small proteins
 TROSY NMR
Transverse Relaxation Optimized Spectroscopy increases
time overwhich NMR signals from neighboring methyl groups can
be detected. The trick is to deuterate the protein then protonate
methyls.

A look into a Proteasome cavity.

This protein is 670 kD! (20S)
Red groups = methyls that are
mobile
Yellow groups = active siteprotein
degradation.

C+E News Feb 5, 2007

 Here Is How It Worked

Nature 445:618 Feb 8, 2007

 Proteasome Function

Core
Proteasome

Ubiquitin Binding Sites top and bottom

 Ubiquitin Targeting a
Cytoplasmic Protein

4th Edition: See pages 1075-

1076, Fig 27-41
5th Edition: See pages 1107-
1109, Fig 27-47, 48

Protein AminoTerminal-aa Half-life

stabilizing
M, G, A, S, T, V >20 hrs
destabilizing
I, N, Y, D, P, L, F, K, R 30 2 min
 Whats New: Free-Electron Lasers
X-ray pulses,
in series of
femtoseconds
on drops
containing
microcrystals.

Free-electron
X ray source
at Stanford
only one ($300
million),
Resolution 2.

Schichting, I. May, 2012 Max-Plank Gesellschaft

 Alpha Helix

Stabilized by H-bonding to every 4th Amino Acid

Alpha Helix Stabilized by Dipole Moments and
Hydrogen Bonds Can be Right or Left Handed
 Alpha Helix H-bonding - Stability
H-bonds to every 4th amino acid
So, amino acid-8 in a 15 aa -helix is H-bonded to aa-11 and aa-5.
What about amino acids at the N- and C-terminal ends?

Instability is brought about by:

1. electrostatic repulsion or attraction charged R groups.
2. adjacent bulky R groups.
3. interaction with R groups 3-4 aas up or down the helix.
4. occurrence of G.
5. interaction of aas at C-terminal and N-terminal ends with any
near aa-R group.
 table 4-1

A
How Long is an 80 amino acid alpha helix?

FACTS to KNOW: One turn: 3.6 aas, 5.4 long

NOW DO IT
How Long is an 80 amino acid alpha helix?

FACTS to KNOW: One turn: 3.6 aas, 5.4 long

80 aas / (3.6 aas/turn) = 22.2 turns

22.2 turns x 5.4 /turn = 120 long

 Sheets
The planarity of the peptide bond and tetrahedral
geometry of the -carbon create a pleated sheet-
like structure
Sheet-like arrangement of backbone is held
together by hydrogen bonds between the backbone
amides in different strands
Side chains protrude from the sheet alternating in
up and down direction
Beta (Pleated Sheet) Structure
Antiparallel Beta-turns
 Antiparallel Beta-turns with Proline

Normal = trans In Beta-turns, Proline is cis

Ramachandran Plot showing 2o Structures
Ramachandran Plot of Pyruvate Kinase
Excludes Glycines they are too flexible
Frequency of Amino Acids in 2o Structure
 Circular Dichroism Spectra

Difference in
right handed
and left
handed
polarized
light on the
extinction
coefficient.

A
 Protein Tertiary Structure
Tertiary structure refers to the overall spatial
arrangement of atoms in a protein

Stabilized by numerous weak interactions between

amino acid side chains.
Largely hydrophobic and polar interactions
Can be stabilized by disulfide bonds

Interacting amino acids are not necessarily next to

each other in the primary sequence.

Two major classes

Fibrous and globular (water or lipid soluble)
 Fibrous Proteins:
From Structure to Function
Alpha Keratin Structure Almost All Alpha Helix
Biochemists at the Hairdressers

Why a Permanent is not a Temporary!

 Chicken Feather -keratin

Carbonizing (under O2 free environment) makes the fiber into a graphite-

like material = light weight, high strength, low cost polymer)
Silk Fibroin is almost all Beta Structure
 Silk

A
Spinnerets of a Spider SEM, Artificially Colored
 Things to Know and Do Before Class
1. The peptide bond and why it is planar.

2. Rotation around the alpha carbon, Ramachandran Plot.

3. Basic idea of X-ray crystallography and how it is used to get

3-D structure.

4. Alpha helix and B-structure.

5. Circular Dichroism spectra: what they demonstrate.

6. Fibrous proteins that are essentially all alpha helix or beta

structure.

7. EOC problems 1-4. We will do some in class and a case

study with music.