Protein Structure Elements

Primary to Quaternary Structure

Learning Objectives

• After this lesson you should be able to:

– Define the structural levels of proteins.– Identify regular secondary structure elements.– Identify the structural units of the protein backbone.– Explain why some backbone conformations are favoured

and some are “forbidden” (not found in natural proteins).– Name properties on which the amino acids can be

grouped.– Explain the driving forces behind protein folding related

to the properties of the backbone and the side chains.

Proteins Are Polypeptides• The peptide bond • A polypeptide chain

Structure Levels• Primary structure = Sequence

(of amino acids)

• Secondary Structure = Helix, sheets/strands, bends, loops & turns (all defined by H-bond pattern in backbone)

• Structural Motif = Small, recurrent arrangement of secondary structure, e.g.– Helix-loop-helix– Beta hairpins– EF hand (calcium binding motif)– Many others…

• Tertiary structure = Arrangement of Secondary structure elements within one protein chain

MSSVLLGHIKKLEMGHS…

• Myoglobin

• Haemoglobin

Quaternary Structure

• Assembly of monomers/subunits into protein complex– Backbone-backbone,

backbone-side-chain & side-chain-side-chain interactions:

• Intramolecular vs. intermolecular contacts.

• For ligand binding side chains may or may not contribute. For the latter, mutations have little effect.

A Bit About Protein Folding

How and why proteins fold

Why Fold?

• Hydrophobic collapse– Hydrophobic residues cluster to “escape”

interactions with water.– Polar backbone groups form secondary

structure to satisfy hydrogen bonding donors and acceptors.

– Initially formed structure is in molten globule state (ensemble).

– Molten globule condenses to native fold via transition state

Hydrophobic Core

• Hydrophobic side chains go into the core of the molecule – but the main chain is highly polar.

• The polar groups (C=O and NH) are neutralized through formation of H-bonds.

Myoglobin

Surface Interior

Hydrophobic vs. Hydrophilic

• Globular protein (in solution)

• Membrane protein

Myoglobin Aquaporin

Hydrophobic vs. Hydrophilic

• Globular protein (in solution)

• Membrane protein

Myoglobin Aquaporin

Cross-section Cross-section

From Unfolded to Native State

G = H - T×S-------------------------G: Free (Gibbs)

energyH: Enthalpy

(interactions)S: Entropy

(conformations/states)

E

U

F

T

G

Unfolded state, ensemble

Native fold, one structure

Transition state, one or more narrow ensembles

Protein Stability & Dynamics

• Folded proteins are:

– Only marginally stable (enthalpy and entropy almost balance at physiological temperatures)

• Allows for easy degradation and reuse.• Amyloid exception.

– Dynamic• “Breathing” motions on pico- to nanosecond scale.• Allows substrates/products to enter/leave enzymes.• Allows allosteric regulation of activity.

Amino Acids• Proteins are built from

amino acids

• Amino group and acid group

• Side chain at C

• Chiral, only one enantiomer found in proteins (L-amino acids)

• 20 natural amino acids

N

O

CC

C

C

C

S

Methionine

Amino Acid Properties

• Many features– Charge +/-

• Acidic vs. basic (pKa)

– Polarity (polar/non-polar)• Type, distribution

– Size• Length, weight, volume, surface area

– Type (Aromatic/aliphatic)

Grouping Amino Acids

Livingstone & Barton, CABIOS, 9, 745-756, 1993

A – AlaC – CysD – AspE – GluF – PheG – GlyH – HisI – IleK – LysL – Leu

M – MetN – AsnP – ProQ – GlnR – ArgS – SerT – ThrV – ValW – TrpY - Tyr

The Evolution Way

• Based on Blosum62 matrix

• Measure of evolutionary substitution probability

Backbone Properties

• Amide bond planarity • 2 degrees of rotational freedom per residue

Ramachandran Plot

• Allowed backbone torsion angles in proteins

N

H

Residue

Peptide bond

Torsion Angles

Characteristics of Helices

• Backbone interactions are local

• Aligned peptide units Dipolar moment

N

C

Helix Types

-Sheets

• Multiple strands sheet– Parallel vs. antiparallel– Twist

• Strand interactions are non-local

• Flexibility– Vs. helices– Folding

Antiparallel Parallel

-Sheets

Thioredoxin

-Sheets

Thioredoxin

-Sheets

Thioredoxin

-Sheets

Thioredoxin

Not All -Sheets Are Flat

• Nitrophorin • Thioredoxin

Residue Patterns

• Helices– Helix capping– Amphiphilic residue

patterns

• Sheets– Amphiphilic residue

patterns– Residue preferences at

edges vs. middle

• Special residues– Proline

• Helix breaker

– Glycine• In turns/loops/bends

N

C

Turns, Loops & Bends Revisited

• Between helices and sheets

• On protein surface

• Intrinsically “unstructured” proteins

Summary

• The backbone of polypeptides form regular secondary structures.– Helices, sheets, turns, bends & loops.

• These are the result of local as well as non-local interactions.

• Secondary structure elements are associated with specific residue patterns.

-sheet and -helices

-sheet -helix

1M8NTheoretical Real