Part I : Introduction to Protein Structure

Part I : Introduction to Part I : Introduction to Protein StructureProtein Structure

Mohamed Ramadan HassanMohamed Ramadan HassanManager of Research & Development Manager of Research & Development

LaboratoryLaboratory Quality Control DepartmentQuality Control Department VACSERAVACSERA

OverviewOverview What are the Importance of Protein Structure ?What are the Importance of Protein Structure ?

The Basics of Protein StructureThe Basics of Protein Structure

Levels of Protein StructureLevels of Protein Structure

Classification of Protein StructureClassification of Protein Structure

OverviewOverview What are the Importance of Protein Structure ?What are the Importance of Protein Structure ?

The Basics of Protein StructureThe Basics of Protein Structure

Levels of Protein StructureLevels of Protein Structure

Classification of Protein StructureClassification of Protein Structure

What are the What are the Importance Importance of Protein Structure ?of Protein Structure ? In the factory of living cells, proteins are the In the factory of living cells, proteins are the

workers, performing a variety of biological tasks.workers, performing a variety of biological tasks. Each protein has a particular 3D structure that Each protein has a particular 3D structure that

brings into close proximity residues that are far brings into close proximity residues that are far apart in the amino acid sequence.apart in the amino acid sequence.

“ “ Structure implies Function “Structure implies Function “ Each protein adopts a particular folding pattern Each protein adopts a particular folding pattern

that determines its function.that determines its function. During normal cells life, most newly synthesized During normal cells life, most newly synthesized

proteins fold spontaneously.proteins fold spontaneously.

SequencSequencee

StructurStructuree

FunctioFunctionn

Common Characters of Common Characters of ProteinsProteins

Physical CharactersPhysical Characters Hydrophobic residues tends to be buried inside the Hydrophobic residues tends to be buried inside the

structure.structure. Hydrophilic residues tends to be exposed to the Hydrophilic residues tends to be exposed to the solvent.solvent.

Electrostatic CharactersElectrostatic Characters Hydrogen bonding between Hydrogen bonding between +ve+ve and and –ve–ve Charged Charged

atoms atoms distantly separated, e.g.; distantly separated, e.g.; – – NN and and –– O O atoms which atoms which

help tohelp to stabilize the structure.stabilize the structure. Structural CharactersStructural Characters

Covalent bonding between Covalent bonding between – SH– SH groups in 2 Cysteine groups in 2 Cysteine residuesresidues in two different chains or in the same chain.in two different chains or in the same chain.

Anfinsen’s Thermodynamic Anfinsen’s Thermodynamic HypothesisHypothesis

“ “ The three-dimensional structure of a native The three-dimensional structure of a native protein in its normal physiological protein in its normal physiological

environment is the one in which the environment is the one in which the Gibbs Gibbs free energyfree energy of the whole system is the of the whole system is the lowestlowest

one; that is, that the native conformation is one; that is, that the native conformation is determined by the totality of interatomic determined by the totality of interatomic interactions and hence by the amino acid interactions and hence by the amino acid

sequence, in a given environment. “sequence, in a given environment. “ ---- Anfinsen’s Nobel Lecture, 1972---- Anfinsen’s Nobel Lecture, 1972

OverviewOverview Why Protein Structure ?Why Protein Structure ?

The Basics of Protein StructureThe Basics of Protein Structure

Levels of Protein StructureLevels of Protein Structure

Classification of Protein StructureClassification of Protein Structure

The Basics of Protein The Basics of Protein StructureStructure

Proteins are linear heteropolymers.Proteins are linear heteropolymers. Contains one or more polypeptide chains.Contains one or more polypeptide chains. Repeat units are 20 natural amino acids.Repeat units are 20 natural amino acids. Total Number of Amino acids from few 10s - 1000s.Total Number of Amino acids from few 10s - 1000s. Proteins enormously varied in 3D shapes ( Proteins enormously varied in 3D shapes ( ““ folds folds ”” ) ) in order to perform their biological activity.in order to perform their biological activity. L-amino acids are the naturally occurring L-amino acids are the naturally occurring

configuration in living organisms.configuration in living organisms.

Common Structure of L-Amino Common Structure of L-Amino AcidAcid

R

C αCN H

HH

H +O

O

AminoAmino

Side Chain = H , CHSide Chain = H , CH3 3 , , ….….

CarboxylateCarboxylate

-Atom lost Atom lost During Peptide During Peptide bond formationbond formation

--------------Atom lost Atom lost duringduringPeptide bond Peptide bond formationformation

CC is a chiral center : i.e.; is a chiral center : i.e.;Has 4 chemically different Has 4 chemically different groups attached to it.groups attached to it. ------------------

-------------------------------------------------------- --------------------------------------

BackboneBackbone

-------------------------

------------------------------------------

-----------------

---------------

------------------

------

------

---

------

------

------

---

---

-----

-----

Aliphatic residues

Alanine

Ala or A

Valine

Val or VOnly heavy atoms are usually shown ( i.e.; no hydrogens Only heavy atoms are usually shown ( i.e.; no hydrogens atoms ).atoms ).Also, residues lacks one oxygen atom in the carboxylate Also, residues lacks one oxygen atom in the carboxylate group.group.

Leucine

Leuor L

Hydrocarbon side chainsHydrocarbon side chains

Aromatic residues

Charged residues

These contain side chains These contain side chains thatthatare charged under are charged under physiologicalphysiologicalconditions, i.e. pH 7.0:-conditions, i.e. pH 7.0:- Acidic – negative charge.Acidic – negative charge. Basic – positive charge.Basic – positive charge.

Polar residues

The odd couple

Can form cis-Can form cis-Peptide bondsPeptide bonds

Formation of Polypeptide Formation of Polypeptide ChainChain

Backbone torsion angles

OverviewOverview Why Protein Structure ?Why Protein Structure ?

The Basics of Protein StructureThe Basics of Protein Structure

Levels of Protein StructureLevels of Protein Structure

Classification of Protein StructureClassification of Protein Structure

This is simply the order of covalent linkages along This is simply the order of covalent linkages along the polypeptide chain, i.e.; the sequence itself.the polypeptide chain, i.e.; the sequence itself.

MMHHGGYYRRTTPPRRSSKKTTDDYYGGCCQQIILLEETTRRAASS

Levels of Protein Levels of Protein StructureStructure

Primary StructurePrimary Structure

Zero StructureZero Structure Amino acid composition, i.e.; percentage of each Amino acid composition, i.e.; percentage of each

single amino acid which can be translated to single amino acid which can be translated to number of each one ( no structural information ).number of each one ( no structural information ).

Levels of Protein Levels of Protein StructureStructure

Local organization of protein backbone:- Local organization of protein backbone:- e.g.;e.g.; αα-helix, -helix, ββ-strand (which assemble into -strand (which assemble into ββ-sheet), -sheet), turn and interconnecting loopturn and interconnecting loop..

Secondary Secondary StructureStructure

Secondary Secondary StructureStructureThe The αα--

helixhelix MyoglobinMyoglobin is the first structure is the first structure

predicted (Pauling, Corey, predicted (Pauling, Corey, Branson 1951) and experimentally Branson 1951) and experimentally solved (Kendrew solved (Kendrew et. al. et. al. 1958).1958).

It is one of the most closely It is one of the most closely packed arrangement of residues.packed arrangement of residues.

Turn: 3.6 residues.Turn: 3.6 residues. Pitch: 5.4 Pitch: 5.4 Å/turn.Å/turn. Rise: 1.5 Å/residue.Rise: 1.5 Å/residue. Dipole: start +ve and end –ve.Dipole: start +ve and end –ve.

Secondary Secondary StructureStructure

Properties of the Properties of the αα--helixhelix Side chains project outwards: Side chains project outwards:

proline only fits the start.proline only fits the start. Amphipathicity if solvent exposed: Amphipathicity if solvent exposed:

hydrophilic residues in cyan; hydrophilic residues in cyan; hydrophobic residues in magenta.hydrophobic residues in magenta.

Secondary Secondary StructureStructure

The The ββ--sheetsheet Side chains project Side chains project

alternately up or down.alternately up or down. Backbone almost fully Backbone almost fully

extended: thus one of extended: thus one of the most loosely packed the most loosely packed arrangements of residues.arrangements of residues.

Secondary Secondary StructureStructure

Topologies of Topologies of ββ-sheets-sheets

Levels of Protein Levels of Protein StructureStructure

Tertiary Tertiary StructureStructure Packing of secondary structure Packing of secondary structure

elements into a compact elements into a compact spatial unit.spatial unit.

““FoldFold”” or domain this is the or domain this is the level to which structure level to which structure prediction is currently possible.prediction is currently possible.

Driving forces in protein Driving forces in protein foldingfolding

Stabilization by forming hydrogen bonds.Stabilization by forming hydrogen bonds. Exposing hydrophilic residues ( charged and Exposing hydrophilic residues ( charged and

polar side chains ) and burying hydrophobic polar side chains ) and burying hydrophobic residues ( aliphatic and aromatic side chains ).residues ( aliphatic and aromatic side chains ).

For small proteins ( usually For small proteins ( usually < 75 residues ).< 75 residues ). Formation of disulfide bridges.Formation of disulfide bridges. Interactions with metal ions.Interactions with metal ions.

The disulfide bondThe disulfide bond It equals disulfide bridges.It equals disulfide bridges. Mostly in extracellular Mostly in extracellular

proteins.proteins. Formed by oxidation of the Formed by oxidation of the

SH (thiol) group of cysteine SH (thiol) group of cysteine residues.residues.

Covalent bond between the Covalent bond between the SSγγ (or (or ‘‘SGSG’’) atoms of two ) atoms of two cysteine residues.cysteine residues.

Levels of Protein Levels of Protein StructureStructure

Quaternary Quaternary StructureStructure Assembly of homo- or Assembly of homo- or

heteromeric protein chains.heteromeric protein chains. Usually the functional unit Usually the functional unit

of a protein, especially for of a protein, especially for enzymes.enzymes.

OverviewOverview Why Protein Structure ?Why Protein Structure ?

The Basics of Protein StructureThe Basics of Protein Structure

Levels of Protein StructureLevels of Protein Structure

Classification of Protein StructureClassification of Protein Structure

Classification of Protein Classification of Protein StructureStructureAll-All- (helical)(helical)

All-All-ββ (sheet)(sheet)

Classification of Protein Classification of Protein StructureStructure

//ββ (parallel (parallel ββ--sheetsheet))

++ββ (antiparallel (antiparallel ββ--sheetsheet))

Most popular Most popular classclass

What is meant by “Domain” Structure

A domain is a compact folding unit of protein A domain is a compact folding unit of protein structure, usually associated with a function.structure, usually associated with a function.

It is usually a It is usually a ““foldfold”” - in the case of - in the case of monomeric soluble proteins.monomeric soluble proteins.

Comprises normally only one protein chain: Comprises normally only one protein chain: rare examples involving 2 chains are known.rare examples involving 2 chains are known.

Domains can be shared between different Domains can be shared between different proteins.proteins.

Homologous FoldsHomologous Folds Hemoglobin and erythrocruorin:

31% sequence identity. Normally at least 25% sequence

identity.. Identical or closely related

functions.

Analogous FoldsAnalogous Folds Hemoglobin and phycocyanin:

9% sequence identity. Structural architechture quite

similar.. Function not conserved.

(I) Structural Comparison (I) Structural Comparison FactsFacts

Proteins adopt only a limited number of folds.Proteins adopt only a limited number of folds. Homologous sequences show very similar Homologous sequences show very similar

structures: variations are mainly in non-structures: variations are mainly in non-conserved regions.conserved regions.

In the absence of sequence homology, some In the absence of sequence homology, some folds are preferred by vastly different folds are preferred by vastly different sequences.sequences.

There are striking regularities in the way in There are striking regularities in the way in which secondary structures are assembled which secondary structures are assembled ( Levitt & Chothia , 1976 ).( Levitt & Chothia , 1976 ).

(II) Structural Comparison (II) Structural Comparison FactsFacts

The The ““active siteactive site”” (a collection of functionally (a collection of functionally critical critical residues) is remarkably conserved, even when the residues) is remarkably conserved, even when the protein fold is differentprotein fold is different..

Structural models (especially those based on homology) Structural models (especially those based on homology) provides insights into possible function for new proteins.provides insights into possible function for new proteins.

Implications for that Implications for that :-:- Protein engineering.Protein engineering.

Ligand/Drug design.Ligand/Drug design. Function assignment for genomic Function assignment for genomic

data.data.