Protein Evolution

Protein Evolution

1. Common supersecondary motifs

   • Helix-loop-helix
   • Coiled-coil
   • Helix bundle
   • Beta-alpha-beta unit
   • Hairpin
   • Beta-meander
   • Greek key
   • Beta-sandwich
   • Combine to form domains
     • Independent folding units in tertiary structure
       • Individual domains have specific function
         • The major driving force in their folding is hydrophobic interactions
     • Important evolutionary units
       • ~40% of known structures in PDB are multidomain proteins
         • Of these >40% contain discontinuous domains
           • Domain insertion is a common evolutionary mechanism
       • 60-80% of genes in genomes code for multidomain proteins
     • Combine with different partners
       • Extend functional repertoire

2. Comparing proteins

   • Basic scoring system
     • Sequence identity % = number of identical residues/number of residues aligned x 100
     • >35% identity means proteins are evolutionarily related
   • Superimposition
   • Root mean square deviation
     • Compares the proximity of residues to one another
       • < 3.5A if related
       • Make alignment
         • Sequence
         • Structure
         • Calculate distance between C-alpha of each pair of aligned residues
           • Pythagoras in 3D
             • Add together for all residues
               • Divide by number of residues
   • So - if 2 proteins have 99% identity...
     • They will have a common 3D structure as long as <50 residues have this
     • Their function is probably similar but even one residue different can completely change function.

3. Recognising domains

   • DNA sequence
     • Limited use
       • Very closely related proteins wil have very similar DNA sequences
       • Perhaps useful for very short evolutionary distances
   • Amino acid sequences
     • Domains have similar amino acid sequences
     • As they diverge the sequence pattern is lost
   • Protein structure
     • Domains have the same fold
       • Has it been seen before?
     • Independently folding units
       • Many computer programs that try to use structural data to identify domains
   • Each domain takes a specific topology/fold
     • There is a limit to how many folds are possible in nature
       • ~10^3-4
         • Even though there are millions of protein sequences there are only so many different ways that structures can be fitted together!

4. Why structural similarity?

   • Divergent evolution from common ancestor
     • If something works it is unlikely to be selected against
     • Structure is much more highly conserved than sequence
       • Makes sense; If a sequence is different that is absolute. But some amino acids have common properties and hence the structure can be retained even when the sequence is not.
   • Convergent evolution
     • Only so many ways to pack helices and strands in 3D space
     • Energetically favourable

5. Protein classification

   • Domains are an important part of structure
     • Structure is conserved because it determines function
       • Protein data bank
   • CATH
     • Class
       • Assigned automatically for >90%
       • Major secondary structure
     • Architecture
       • Gross orientation of secondary structures
         • Shape of the fold
       • Beta-roll
       • Up-down bundle
       • Alphabeta-prism
       • alpha-beta-alpha sandwich
     • Topology
       • Connections
       • Number of 2ry strctures
     • Homologous superfamily
       • Highly similar structures and functions
       • Is there enough evidence for shared evolutionary origin?
     • Process
       • 1. Chop proteins into domains
         • 2. Sequence and structural analysis programs group by evolutionary and structural families
     • CATH domain structures and sequence relatives in sequenced genomes are fairly representative
     • Applications
       • "Ab initio" methods using protein structure
         • Algorithms for recognising boundaries
           • Swindells, 1995 - Detective
             • Each domain should have a recognisable hydrophobic core
           • Siddiqui and Barton, 1995 - DOMAK
             • Residues comprising a domain make more internal contacts than external ones
           • Holm & Sander, 1994 - PUU
             • Parser for protein folding units
               • Maximal interaction within domains
               • Minimal interaction between domains
           • Seek consensus; in practice ~20% of cases
           • E.g. Beta-amylase
             • PUU: 1 domain
             • DOMAK: 2 domains
             • Detective: 3 domains
       • Sequence methods
         • Sequence-sequence comparison
           • BLAST
           • NW
         • Sequence-profile comparison
           • PSI-BLAST
           • Align sequences and colour-code degree of conservation
           • If identity <35%
       • Structural methods
         • Contact map
           • Points of contact between residues in a protein
         • Distance matrices
       • Enter text here
   • Structural classification of proteins
     • Class
     • Fold
     • Superfamily
     • Family
     • Species
   • Protein family database Pfam
     • Superfamily
     • Clade
     • Sequence based
       • Confers with SCOP
   • Methods vary but agree on 60-70% of cases

6. Homology

   • An absolute value
     • Either homologous or not!
   • Orthologs
     • Common ancestor
       • Different species
       • Same function
   • Paralogs
     • Common ancestor
       • Gene duplication
       • Same or different species
       • Different but related function
   • At least 2 conditions must be met:
     • Significant structural similarity
     • Significant sequence similarity
     • Functional similarity
   • e.g. toxins
     • Cholera
     • Pertussis
     • Heat stable enterotoxin
     • High structural similarity
       • Related functions
       • No evidence that they evolved from a common ancestor

7. Rossmann fold

   • NAD binding domain
     • Cofactor that reversibly accepts a hydride ion
       • Lost or gained by the substrate in the redox reaction
       • Found in all living cells
         • Metabolism
           • Accepts or donates electrons in redox
             • Remove two hydrogen atoms from reactant (R)
               • Hydride ion H-
                 • Reduces NAD+ to NADH
               • Proton H+
                 • Released into solution
       • 2 nucleotides joined through their phosphate group
         • One contains an adenine base
         • The other nicotinamide
   • One of the most ubiquitous domains
   • Alpha-beta fold
     • Central beta sheet
       • Surrounded by approximately 5 alpha helices
       • Strands in characteristic order 654123
         • Crossover forms binding site
   • Lactate dehydrogenase
     • Has a Rossman fold at N-terminal domain
     • Convert L-lactate to pyruvate
       • Last step in anaerobic glycolysis
     • C-terminal catalytic domain
       • Substrate specificity
         • Precise reaction
       • Specific to lactate/malate dehydrogenases
         • Malate dehydrogenase
           • Interconversion of malate to oxaloacetate
           • N-terminal domain is a Rossmann fold
           • C-terminal catalytic domain
         • Paralogs
           • 17% sequence identity in humans
             • Duplication event
             • BUT structurally very similar
               • Function of NAD-binding domain conserved
                 • Change in sequence confers change in substrate specificity
     • Human/zebrafish orthologs
       • 76% sequence identity
         • Suggests speciation from common ancestor
   • Alcohol dehydrogenase
     • Two ADCDs flank a Rossmann fold
     • Same structure as LDH
       • 17% identity
   • Combine with different catalytic domains to achieve different functions

8. Sequence diversity

   • Different evolutionary constraints in different positions in the protein structure
     • Surface residues have the least evolutionary constraints and can also accommodate small insertions and deletions
     • Core residues more highly conserved, critical for folding and stability
     • Functional residues also highly conserved
       • Critical for enzyme function or for protein-protein interactions
   • Structural diversity
     • The core is usually highly conserved
       • Within a family ~50%
     • Residue insertions in loops connecting 2ry structures
     • Substitutions can cause shifts in orientations of 2ry structure
     • Domain embellishments
   • Functional diversity
     • Domain superfamily
       • Dependent on the fold
         • Some can support many similar functions
           • P-loop hydrolases
         • Some have limited repertoire of functions
           • Globins
     • 1 amino acid change can change the function of a protein
     • Can share <10% sequence identity but still have the same function in different organisms
     • Defining function
       • Biochemical
         • Conservation
           • Chemistry?
           • Substrate?
           • Product?
       • Biological
         • Is cell localisation conserved?
           • Myoglobin
           • Haemoglobin
       • Schema
         • Enzyme classification system
         • GO terms

9. Challenges in comparing protein structures

   • Insertions or deletions of residues
     • Usually in connecting loops not secondary structures
     • Indels
     • Structural cores usually highly conserved
       • Can still be considerable structural differences between relatives outside the core.
     • Insertions usually in loops connecting secondary structures
     • Substitutions can shift orientations of secondary structures
     • Coping
       • Ignore variable loop regions
         • Only compare secondary structures
       • Use algorithms which explicitly handle insertions/deletions
         • Dynamic programming
         • Simulated annealing

10. CATHEDRAL

    • Combines rapid graph theory secondary structure filter with dynamic programming
      • Accurate residue alignment
      • SVM
        • Combine scores
          • Assess significance of match
      • Scan against all domain structural representatives in CATH
    • Fast structure comparison
      • Dihedral angles + chirality
        • Create overlap graph
          • Largest common structural motif
          • Compare using Bron Kerbosch algorithm
            • Largest common graph
      • Generally ~1000x faster than residue-based methods

11. SSAP

    • Residue-based method
      • Double dynamic programming
    • Compare vector environments
      • Path matrix
        • Generate path for each
          • Compare with dynamic algorithm
            • Add path to summary matrix
              • Apply dynamic algorithm to summary matrix
                • Final alignment