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Review I: Protein StructureReview I: Protein Structure

Rajan Munshi

BBSI @ Pitt 2006BBSI @ Pitt 2006

Department of Computational BiologyUniversity of Pittsburgh School of Medicine

May 23, 2006

Amino Acids

Building blocks of proteins20 amino acidsLinear chain of amino acids form a peptide/proteinα−carbon = central carbonα−carbon = chiral carbon, i.e. mirror images: L and D isomersOnly L isomers found in proteinsGeneral structure:

(R = side chain)

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Amino Acids (contd.)

R group variesThus, can be classified based on R groupGlycine: simplest amino acidSide chain R = HUnique because Gly α carbon is achiral

Glycine, Gly, G

H

CαH2N COOH

H

Amino Acids: Structures

orange = non-polar, hydrophobicneutral (uncharged)

blue = R (side chain)

green = polar, hydrophillic,neutral (uncharged)

magenta = polar, hydrophillic,acidic (- charged)

light blue = polar, hydrophillic,basic (+ charged)

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Amino Acids: ClassificationNon-polar, hydrophobic,neutral (uncharged)Alanine, Ala, AValine, Val, VLeucine, Leu, LIsoleucine, Ile, IProline, Pro, PMethionine, Met, MPhenylalanine, Phe, FTryptophan, Trp, W

Polar, hydrophillic,neutral (uncharged)Glycine, Gly, GSerine, Ser, SThreonine, Thr, TCysteine, Cys, CAsparagine, Asn, NGlutamine, Gln, QTyrosine, Tyr, Y

Polar, hydrophillic,Acidic (negatively charged)Aspartic acid, Asp, DGlutamic acid, Glu, E

Polar, hydrophillic,basic (positively charged)Lysine, Lys, KArginine, Arg, RHistidine, His, H

Peptide Bond Formation

Condensation reactionBetween –NH2 of n residue and –COOH of n+1 residueLoss of 1 water moleculeRigid, inflexible

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Peptides/Proteins

Linear arrangement of n amino acid residues linked by peptide bondsn < 25, generally termed a peptiden > 25, generally termed a proteinPeptides have directionality, i.e. N terminal C-terminal

R1

CαH2N C

H

CαN COOH

HO

R2

nN terminal

C terminal

Peptide bond

Hierarchy of Protein Structure

Four levels of hierarchyPrimary, secondary, tertiary, quarternary

Primary structure: Linear sequence of residuese.g: MSNKLVLVLNCGSSSLKFAV …e.g: MCNTPTYCDLGKAAKDVFNK …

Secondary Structure: Local conformation of thepolypeptide backboneα-helix, β-strand (sheets), turns, other

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Secondary Structure: α-helix

Most abundant; ~35% of residues in a proteinRepetitive secondary structure3.6 residues per turn; pitch (rise per turn) = 5.4 ÅC′=O of i forms H bonds with NH of residue i+4Intra-strand H bondingC′=O groups are parallel to the axis; side chains pointaway from the axisAll NH and C′O are H-bonded, except first NH and last C′OHence, polar ends; present at surfacesAmphipathic

α-helix (contd.)

N terminal

C terminal

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α−helix Variations

Chain is more loosely or tightly coiled310-helix: very tightly packedπ−helix: very loosely packedBoth structures occur rarely Occur only at the ends or as single turns

Hemoglobin (PDB 1A3N)

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β−sheets

Other major structural elementBasic unit is a β-strandUsually 5-10 residuesCan be parallel or anti-parallel based on therelative directions of interacting β-strands“Pleated” appearance

β−sheetsUnlike α-helices:Are formed with different parts of the sequenceH-bonding is inter-strand (opposed to intra-strand)Side chains from adjacent residues are onopposite sides of the sheet and do not interactwith one another

Like α-helices:Repeating secondary structure (2 residues per turn)Can be amphipathic

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Parallel β−sheets

The aligned amino acids in the β-strand all run in the same biochemical direction, N- to C-terminal

Anti-parallel β−sheets

The amino acids in successive strands have alternating directions, N-terminal to C-terminal

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Nucleoplasmin (PDB 1K5J)

Amino Acid Preferences (1)α-helix formingThe amino acid side chain should cover and protect the backbone H-bonds in the core of the helixAla, Leu, Met, Glu, Arg, Lys: good helix formersPro, Gly, Tyr, Ser: very poor helix formers

β-strand formingAmino acids with large bulky side chains prefer to formβ-sheet structuresTyr, Trp, Ile, Val, Thr, Cys, Phe

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Amino Acid Preferences (2)

Secondary structure disruptorsGly: side chain too smallPro: side chain linked to α-N, has no N-H toH-bond; rigid structure due to ringAsp, Asn, Ser: H-bonding side chains competedirectly with backbone H-bonds

Turns/Loops

Third "classical" secondary structureReverses the direction of the polypeptide chain Located primarily on protein surfaceContain polar and charged residuesThree types: I, II, III

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Phosphofructokinase (PDB 4PFK)

The Torsional Angles: φ and ψ

Each amino acid in a peptide has two degreesof backbone freedomThese are defined by the φ and ψ anglesφ = angle between Cα―Nψ = angle between Cα―C’

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The Ramachandran PlotPlot of allowable φ and ψ anglesφ and ψ refer to rotations of two rigid peptide units around Cα

Most combinations produce steric collisionsDisallowed regions generallyinvolve steric hindrance between side chain Cβ methylene group and main chain atoms

The Ramachandran Plot (contd.)

Theoreticallypossible;

energeticallyunstable

π-helix

310-helix

Anti-parallelβ-sheet

Parallelβ-sheet

White: sterically disallowed(except Gly)

Red: no steric clashesYellow: “allowable” steric

clashes

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Protein Motifs

(Simple) combinations of secondary structure elements with a specific geometric arrangement“Super-secondary structures”Can be associated with a specific function, e.g. DNA binding, or metal ion bindingCan be part of a larger functional and/or structural assembly (“domain”)

Helix-Turn-Helix (HTH)

Simplest α-helix motif: 2 α helices joined by a loopAlso called Helix-Loop-Helix, HLHCommon structural motif forDNA binding proteinsOne helix recognizes specificsequence of nucleotides and fitsinto the groove in the DNA doublehelix; the other stabilizes the boundconfiguration

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EF Hand

Specific for several differentcalcium-binding proteinsE.g. calmodulin, Troponin-C

Hairpin β-motif

Also called β-hairpin2 adjacent anti-parallel strandsjoined by a loop

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Greek Key motif

4 adjacent anti-parallel β-strands

β-α-β Fold

Parallel β-strands connect by an α helixFound in most proteins with parallel β strands. E.g. Triosephosphate isomerase

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Rossman Fold

Unique example of a β−α−β fold3 parallel β-sheets with 2 linkingα helicesOften seen in nucleotide-bindingproteins

Domains

Primary structureSecondary structureSuper-secondary structureDomainsFundamental unit of tertiary structure(Part of a) polypeptide chain that can fold independently into a stable tertiary structure E.g. catalytic domain of protein kinase; binding pocket of a ligand

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N-terminal

C-terminal

phosphate binding

loop

catalyticloop

3D Structure of a Protein Kinase Domain

Quarternary Structure

Spatial organization of subunits to form functional proteinE.g. Hemoglobin2 α chains, 2 β chainsEach chain binds heme (Fe)Forms an α2β2 tetramer

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Putting it all together

Primary Structure… KAAWGKVGAHA …

Secondary Structureα-helix, β-sheets, turns/loops

Tertiary Structurea single chain (α, β)

Quarternary Structureα2β2

Super-secondary StructureHeme-binding pocket/domain (His)

Additional Reading

General informationBiochemistry, 5th ed., Berg, Tymoczko, StryerBiochemistry, 3rd ed., Voet & Voet

Detailed informationProteins, 2nd ed., CreightonIntroduction to Protein Structure, 2nd ed., Branden & Tooze

InternetImages: Protein Data Bank (PDB): www.rcsb.org/pdbNumerous wesbites (Google protein secondary structure)