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Principles of Protein Structure
primary structure
ACDEFGHIKLMNPQRSTVWY
Different Levels of Protein Structure
NH2
Lysine
Histidine
Valine
Arginine
Alanine
COOH
Common Secondary Structure Elements
• The Alpha Helix
Properties of alpha helix
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3.6 residues per turn, 13 atoms between H-bond donor and acceptor
approx. -60º;  approx. -40º
H- bond between C=O of ith residue & -NH of (i+4)th residue
First -NH and last C=O groups at the ends of helices do not participate in Hbond
Ends of helices are polar, and almost always at surfaces of proteins
Always right- handed
Macro- dipole
Alpha Helix
Helical wheel
Residues i, i+4, i+7 occur on
one face of helices, and
hence show definite pattern
of hydrophobicity/
hydrophilicity
Association of helices: coiled coils
Introduction
to
Molecular
Biophysics
These coiled coils have a heptad repeat abcdefg with nonpolar residues at
position a and d and an electrostatic interaction between residues e and g.
Isolated alpha helices are
unstable in solution but are
very stable in coiled coil
structures because of the
interactions between them
The chains in a coiled-coil have
the polypeptide chains aligned
parallel and in exact axial
register. This maximizes
coil formation between chains.
The coiled coil is a protein motif that is often used to control oligomerization.
They involve a number of alpha-helices wound around each other in a highly
organised manner, similar to the strands of a rope.
The Leucine Zipper Coiled Coil
Introduction to Molecular Biophysics
Initially identified as a structural motif in proteins involved in eukaryotic
transcription. (Landschultz et al., Science 240: 1759-1763 (1988).
Originally identified in the liver transcription factor C/EBP which has a Leu
at every seventh position in a 28 residue segment.
Association of helices: coiled coils
The helices do not have to run in the same direction for this type of
interaction to occur, although parallel conformation is more common.
Antiparallel conformation is very rare in trimers and unknown in
pentamers, but more common in intramolecular dimers, where the two
helices are often connected by a short loop.
Chan et al., Cell 89, Pages 263-273.
Basis for the helical dipole
In an alpha helix all of the peptide
dipoles are oriented along the
same direction.
Consequently, the alpha helix has
a net dipole moment.
Since the dipole moment of a peptide bond is 3.5 Debye units, the alpha
helix has a net macrodipole of:
n X 3.5 Debye units (where n= number of residues)
This is equivalent to 0.5 – 0.7 unit charge at the end of the helix.
The amino terminus of an alpha helix is positive and the
carboxy terminus is negative.
Structure of human TIM
Two helix dipoles are seen to play
important roles:
1.
2.
Stabilization of inhibitor 2-PG
Modulation of pKa of active site
His-95.
Helical Propensities
Ala
Arg
Lys
Leu
Met
Trp
Phe
Ser
Gln
Glu
Cys
Ile
Tyr
Asp
Val
Thr
Asn
His
Gly
Pro
-0.77
-0.68
-0.65
-0.62
-0.50
-0.45
-0.41
-0.35
-0.33
-0.27
-0.23
-0.23
-0.17
-0.15
-0.14
-0.11
-0.07
-0.06
0
~3
Common Secondary Structure Elements
• The Beta Sheet
Secondary structure: reverse turns
Secondary Structure:
Phi & Psi Angles Defined
• Rotational constraints emerge from interactions with bulky
groups (ie. side chains).
• Phi & Psi angles define the secondary structure adopted by
a protein.
The dihedral angles at C atom of every residue
provide polypeptides requisite conformational
diversity, whereby the polypeptide chain can fold into
a globular shape
Ramachandran Plot
Secondary Structure
Table 10
Phi & Psi angles for Regular Secondary
Structure Conformations
Structure
Antiparallel b-sheet
Parallel b-Sheet
Right-handed -helix
310 helix
p helix
Polyproline I
Polyproline II
Polyglycine II
Phi (F)
-139
-119
+64
-49
-57
-83
-78
-80
Psi(Y)
+135
+113
+40
-26
-70
+158
+149
+150
Beyond Secondary Structure
Supersecondary structure (motifs): small, discrete, commonly
observed aggregates of secondary structures
 b sheet
 helix-loop-helix
 bb
Domains: independent units of structure
 b barrel
 four-helix bundle
*Domains and motifs sometimes interchanged*
Common motifs
Supersecondary structure:
Crossovers in b--b-motifs
Left handed
Right handed
EF Hand
• Consists of two perpendicular 10 to 12 residue alpha helices with
a 12-residue loop region between
• Form a single calcium-binding site (helix-loop-helix).
• Calcium ions interact with residues contained within the loop
region.
• Each of the 12 residues in the loop region is important for
calcium coordination.
• In most EF-hand proteins the residue at position 12 is a
glutamate. The glutamate contributes both its side-chain oxygens
for calcium coordination.
Calmodulin, recoverin : Regulatory proteins
Calbindin, parvalbumin: Structural proteins
EF Fold
Found in Calcium binding proteins such as Calmodulin
Helix Turn Helix Motif
•Consists of two  helices and a short extended amino acid chain between them.
•Carboxyl-terminal helix fits into the major groove of DNA.
•This motif is found in DNA-binding proteins, including l repressor, tryptophan
repressor, catabolite activator protein (CAP)
Leucine Zipper
Rossman Fold
•The beta-alpha-beta-alpha-beta subunit
•Often present in nucleotide-binding proteins
What is a Protein Fold?
 Compact, globular folding arrangement of the polypeptide chain
 Chain folds to optimise packing of the hydrophobic residues in the interior
core of the protein
Common folds
Tertiary structure examples: All-
Cytochrome C
four-helix bundle
Alamethicin
The lone helix
Rop
helix-turn-helix
Tertiary structure examples: All-b
b sandwich
b barrel
Tertiary structure examples: /b
placental ribonuclease
inhibitor
/b horseshoe
triose phosphate
isomerase
/b barrel
Four helix bundle
•24 amino acid peptide with a hydrophobic surface
•Assembles into 4 helix bundle through hydrophobic regions
•Maintains solubility of membrane proteins
Oligonucleotide Binding (OB) fold
TIM Barrel
•The eight-stranded  /b barrel (TIM barrel)
•The most common tertiary fold observed in
high resolution protein crystal structures
•10% of all known enzymes have this domain
Zinc Finger Motif
Domains are independently folding structural units.
Often, but not necessarily, they are contiguous on the peptide chain.
Often domain boundaries are also intron boundaries.
Domain swapping:
Parts of a peptide chain can reach into neighboring
structural elements: helices/strands in other domains or
whole domains in other subunits.
Domain swapped diphteria toxin:
Transmembrane Motifs
• Helix bundles
Long stretches of apolar amino acids
Fold into transmembrane alpha-helices
“Positive-inside rule”
Cell surface receptors
Ion channels
Active and passive transporters
• Beta-barrel
Anti-parallel sheets rolled into cylinder
Outer membrane of Gram-negative bacteria
Porins (passive, selective diffusion)
Quaternary Structure
• Refers to the organization of subunits in a protein with multiple subunits
• Subunits may be identical or different
• Subunits have a defined stoichiometry and arrangement
• Subunits held together by weak, noncovalent interactions (hydrophobic,
electrostatic)
• Associate to form dimers, trimers, tetramers etc. (oligomer)
• Typical Kd for two subunits: 10-8 to 10-16M (tight association)
–Entropy loss due to association - unfavorable
–Entropy gain due to burying of hydrophobic groups - very favourable
Structural and functional advantages of
quaternary structure
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•
•
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Stability: reduction of surface to volume ratio
Genetic economy and efficiency
Bringing catalytic sites together
Cooperativity (allostery)
Quaternary structure of
multidomain proteins