1. Introduction to Protein Structure
Paolo Marcatili

2. Learning Objectives Outline the basic levels of protein structure.
Evaluate the quality of protein structures
Navigate a protein structure using the program PyMOL.
Generate publication-ready protein figures

3. Learning Objectives Predict protein features from its sequence
Build a model by homology and predict its accuracy
Refine your model
Predict the effects of mutations on stability and binding affinity

4. Activity - Predict the structure of a pathogenic protein
- Predict the role of mutations in antibody affinity

5. Levels of Protein Structure
Primary to Quaternary Structure

6. Amino Acids Proteins are built from amino acids
Amino group and acid group
Side chain at Ca
Chiral, only one enantiomer found in proteins (L-amino acids)
20 natural amino acids Ce Sd Cg Cb C Ca N O
Methionine

7. The Amino Acids Thr (T) Phe (F) Val (V) Ala (A) His (H) Arg (R)
Ser (S)
Leu (L)
Cys (C)
Met (M)
Asp (D)
Lys (K)
Asn (N)
Ile (I)
Trp (W)
Gln (Q)
Glu (E)
Tyr (Y)
Pro (P)
Gly (G)

8. How to Group Them? Many features Charge +/- Polarity (polar/non-polar)
Acidic vs. basic (pKa)
Polarity (polar/non-polar)
Type, distribution
Size
Length, weight, volume, surface area
Type (Aromatic/aliphatic)

9. Grouping Amino Acids Acid deriv.

10. The Simple Aliphatic
Val (V)
Ala (A)
Leu (L)
Met (M)
Ile (I)

11. The Small Polar
Ser (S)
Cys (C)
Cysteine
Thr (T)
Cystine

12. The Unusual P, G Also aliphatic Structural impact
Strictly speaking, proline is an imino acid
Gly (G)
Pro (P)

13. The Acidic and Their Derivatives
Asp (D)
Asn (N)
Glu (E)
Gln (Q)

14. The Basic
Arg (R)
Lys (K)
His (H)

15. The Aromatic
Phe (F)
His (H)
Trp (W)
Tyr (Y)

16. Hypothesis i) Structure <=> Function
ii) A protein has a single stable structure
iii) This structure is determined by the sequence alone
iv) The protein can reach this fold autonomously (Anfinsen)

17. Hypothesis
i) Structure <=> Function convergent and divergent evolution
ii) A protein has a single stable structure
Metastability, serpins, prions
iii) This structure is determined by the sequence alone
PTMs, epigenetics
iv) The protein can reach this fold autonomously (Anfinsen)
Enzymes, Chaperones, boiled eggs

18. Native State
The proteins reach the conformation in which they minimize their Free Energy

19. Native State
The proteins reach the conformation in which they minimize their Free Energy
They just visit random conformations
until they find the best one?

20. Native State
The proteins reach the conformation in which they minimize their Free Energy
They just visit random conformations
until they find the best one?
NO!
it would take more than the age of universe (Levinthal's Paradox)

21. Folding Funnel

22. Proteins Are Polypeptides
The peptide bond
A polypeptide chain

23. Ramachandran Plot
Allowed backbone torsion angles in proteins N H

24. 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…

25. 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.
Myoglobin
Haemoglobin a a b

26. 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

27. Hydrophobic vs. Hydrophilic
Globular protein (in solution)
Membrane protein (in membrane)
Myoglobin
Aquaporin

28. Hydrophobic vs. Hydrophilic
Globular protein (in solution)
Membrane protein (in membrane)
Cross-section
Cross-section
Myoglobin
Aquaporin

29. Helix Types

30. b-Sheets Multiple strands  sheet Flexibility
Parallel vs. antiparallel
Twist
Flexibility
Vs. helices
Folding
Structure propagation (amyloids)
Other…
Thioredoxin

31. b-Sheets Multiple strands  sheet Flexibility
Parallel vs. antiparallel
Twist
Flexibility
Vs. helices
Folding
Structure propagation (amyloids)
Other…

32. b-Sheets Multiple strands  sheet Flexibility
Parallel vs. antiparallel
Twist
Flexibility
Vs. helices
Folding
Structure propagation (amyloids)
Other…

33. b-Sheets Multiple strands  sheet Flexibility
Parallel vs. antiparallel
Twist
Flexibility
Vs. helices
Folding
Structure propagation (amyloids)
Other…

34. b-Sheets Multiple strands  sheet Strand interactions are non-local
Parallel vs. antiparallel
Twist
Strand interactions are non-local
Flexibility
Vs. helices
Folding
Antiparallel
Parallel

35. Turns, Loops & Bends Between helices and sheets On protein surface
Intrinsically “unstructured” proteins

36. 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.

37. evolution, structure or function… ?

Domain Definition(s)
From wikipedia
protein domain is a part of protein sequence and structure
that can evolve, function, and exist independently of the rest
of the protein chain.
evolution, structure or function… ?

38. evolution, structure or function… ?

Domain Definition(s)
From wikipedia
protein domain is a part of protein sequence and structure
that can evolve, function, and exist independently of the rest
of the protein chain.
evolution, structure or function… ?

Working definition:
A structural domain is a compact, globular sub-structure
with more interactions within it than with the rest of the protein.

39. SCOP & CATH Similar definitions, but
SCOP ~ human (good accuracy, less systematic, less data)
CATH ~ automatic (more data, more systematic)

40. SCOP All alpha proteins (151) All beta proteins (111)
Alpha and beta proteins (a/b) (117) Mainly parallel beta sheets (beta-alpha-beta units)
Alpha and beta proteins (a+b) (212) Mainly antiparallel beta sheets (segregated alpha and beta regions)

41. Fold: Major structural similarity
Proteins are defined as having a common fold if they have
the same major secondary structures in the same arrangement and
with the same topological connections.
Different proteins with the same fold often have peripheral elements
of secondary structure and turn regions that differ in size and conformation.
In some cases, these differing peripheral regions may comprise
half the structure. Proteins placed together in the same fold category
may not have a common evolutionary origin:
the structural similarities could arise
just from the physics and chemistry of proteins
favoring certain packing arrangements and chain topologies.

42. Superfamily: Probable common evolutionary origin
Proteins that have low sequence identities,
but whose structural and functional features suggest
that a common evolutionary origin is probable
are placed together in superfamilies.
For example, actin, the ATPase domain
of the heat shock protein, and hexokinase together form
a superfamily.

43. Family: Clear evolutionarily relationship
Proteins clustered together into families are clearly
evolutionarily related. Generally, this means that
pairwise residue identities between the proteins
are 30% and greater. However, in some cases
similar functions and structures provide definitive evidence
of common descent in the absense of high sequence identity;
for example, many globins form a family though some members
have sequence identities of only 15%.

44. Some Folds

45. CATH hierarchy
Class
Architecture
Topology Homologous superfamily

46. Pfam If we dont have structure, how can we know
domain arrangment or protein family
of our target?

47. How to detect domains in a protein?
SCOP / CATH
PFAM
transmembrane regions
Blocks in the MSA
low-complexity regions

48. Protein Data Bank http://www.rcsb.org/ Contents File structure
Types of structures
Structure reports & summaries
Quality check
Searching
Molecule of the Month

49. PDB Growth

50. PDB File Fields
COLUMNS DATA TYPE FIELD DEFINITION – 6 Record name "ATOM" Integer serial Atom serial number. 13 – 16 Atom name Atom name. 17 Character altLoc Alternate location indicator. 18 – 20 Residue name resName Residue name. 22 Character chainid Chain identifier. 23 – 26 Integer resSeq Residue sequence number. 27 AChar iCode Code for insertion of residues. 31 – 38 Real(8.3) x Orthogonal coordinates for X in Angstroms 39 – 46 Real(8.3) y Orthogonal coordinates for Y in Angstroms 47 – 54 Real(8.3) z Orthogonal coordinates for Z in Angstroms 55 – 60 Real(6.2) occupancy Occupancy. 61 – 66 Real(6.2) tempFactor Temperature factor. 77 – 78 LString(2) element Element symbol, right-justified. 79 – 80 LString(2) charge Charge on the atom.