Evolution of protein coding sequences

1. Evolution of protein coding sequences

2. Single substitution Multiple substitution Coincidental substitution C A T G A A A A C C 1 change, 1 difference 2 changes, 1 difference 2 change, 1 difference Parallel substitution Convergent substitution Back substitution C C T A A A A A C T C 2 changes, no difference 3 changes, no difference 2 changes, no difference Kinds of nucleotide substitutions Given 2 nucleotide sequences, how their similarities and differences arose from a common ancestor? We assume A the common ancestor:

3. Important properties inherent to the standard genetic code

4. Synonymous vs nonsynonymous substitutions • Nondegenerate sites: are codon position where mutations always result in amino acid substitutions. (exp. TTT (Phenylalanyne, CTT (leucine), ATT (Isoleucine), and GTT (Valine)). • Twofold degenerate sites: are codon positions where 2 different nucleotides result in the translation of the same aa, and the 2 others code for a different aa. (exp. GAT and GAC code for Aspartic acid (asp, D), whereas GAA and GAG both code for Glutamic acid (glu, E)). • Threefold degenerate sites: are codon positions where changing 3 of the 4 nucleotides has no effect on the aa, while changing the fourth possible nucleotide results in a different aa. There is only 1 threefold degenerate site: the 3rd position of an isoleucine codon. ATT, ATC, or ATA all encode isoleucine, but ATG encodes methionine.

5. • Five amino-acids are encoded by 4 codons which differ only in the third position. These sites are called “fourfold degenerate” sites Standard genetic code • Fourfold degenerate sites: are codon positions where changing a nucleotide in any of the 3 alternatives has no effect on the aa. exp. GGT, GGC, GGA, GGG(Glycine); CCT,CCC,CCA,CCG(Proline) • Three amino acids: Arginine, Leucine and Serine are encoded by 6 different codons:

6. Standard genetic code • Nine amino acids are encoded by a pair of codons which differ by a transition substitution at the third position. These sites are called “twofold degenerate” sites. Transition: A/G; C/T • Isoleucine is encoded by three codons(with a threefold degenerate site) • Methionine and Triptophan are encoded by single codon •Three stop codons: TAA, TAG and TGA

7. Evolution of protein coding sequences • Some amino acid substitutions require more DNA substitutions than others • Ile  Thr : at least one DNA change • AUU  ACU • AUC  ACC • AUA  ACA • Ile  Cys: at least two DNA changes • AUU (Ile)  AGU (Ser)  UGU (Cys) • AUU (Ile)  UUU (Phe)  UGU (Cys)

8. GluVal Phe SEQ.1 GAA GTTTTT SEQ.2 GAC GTCGTA AspVal Val TTT(F:Phe) 2 1 TTA(L:Leu) GTT(V:Val) GTA(V:Val) Example: 2 homologous sequences •Codon 1: GAA --> GAC ;1 nuc. diff., 1 nonsynonymous difference; •Codon 2: GTT --> GTC ;1 nuc. diff., 1 synonymous difference; •Codon 3: counting is less straightforward: Path 1 : implies 1 non-synonymous and 1 synonymous substitutions; Path 2 : implies 2 non synonymous substitutions;

9. Evolution of protein coding sequences • Redundancy of the genetic code • Biochemical properties of amino acids • Under neutral evolution (no effect of selection) amino acids should replace each other with a probability determined by the number of DNA substitutions

10. Evolution of protein coding sequences • Some amino acid substitutions require more DNA substitutions than others • Ile  Thr : at least one DNA change • AUU  ACU • AUC  ACC • AUA  ACA • Ile  Cys: at least two DNA changes • AUU (Ile)  AGU (Ser)  UGU (Cys) • AUU (Ile)  UUU (Phe)  UGU (Cys)

11. Rates and patterns of nucleotide substitution • Influenced by three things • Functional constraint (negative selection) • Positive selection • Mutation rate

12. Rate of nucleotide substitution • K = mean number of substitutions per site • T = time since divergence • rate = r = number of substitutions per site per year • r = K/2T Ancestral sequence T T Sequence 1 Sequence 2

13. Gene tree - Species tree • Duplication Time • Duplication Speciation Speciation A C B C A B B C A Species tree Gene tree Genomes 2 edition 2002. T.A. Brown

14. Common ancestor of sequences Ancestral species Allele B Allele A speciation Time Gorilla Human

15. Evolution of protein-coding sequences • The Genetic Code is redundant • Some nucleotide changes do not change the amino acid coded for • 3rd codon position often synonymous • 2nd position never • 1st position sometimes

16. Standard Genetic Code

17. rates • In general ... • Rates of nucleotide substitution are lowest at nondegenerate sites (0.78 x 10-9 per site per year) • Intermediate at two-fold degenerate sites (2.24 x 10-9) • Highest at fourfold degenerate sites (3.71 x 10-9)

18. Effect of amino acid substitutions • Deleterious 86% • Neutral 14% • Advantgageous 0.0% ? (very low) • In protein coding sequences, selection is often acting to remove changes • Less common outcome is drift of neutral changes • Rarely see positive selection for advantageous changes

19. Functional Constraint • Proteins often have some functional constraint • The stronger the functional constraint, the slower the rate of evolution

20. Haemoglobin • Haeme pocket is highly constrained at protein seq. level • Remainder of protein only constrained to be hydrophillic

21. Histone 4 • Two copies in Histone octamer • Forms complex with other histones and binds DNA into chromatin • Almost the whole protein is highly constrained

22. Fibrinopeptides • Hardly any sequence constraint

24. Rates and Patterns • Patterns of change can be informative of the function of a protein • Different genes evolve at different rates • Amino acids that are always conserved are likely to be critical to the function

25. Biochemical properties

27. Histone 4 • Highly conserved protein • Compare human and wheat H4 genes • 55 DNA differences • 2 amino acid differences • Val  Ile (both aliphatic) • Lys  Arg (both charged)

28. Evolution of non-coding regions • homologous sequences • e.g., compare introns of homologous genes • 5’ UTR and 3’ UTR (untranslated region) • Pseudogenes

31. Synonymous substitution rate variation • Synonymous rates may differ between genes • How come? • Maybe different mutation rates in different parts of the genome

32. Variation in the rates of synonymous substitutions: Secondary structure constraints • Stems in secondary RNA structures are more constrained than loops.