Abstract
BACKGROUND: Protein evolution is particularly shaped by the conservation of the amino acids' physico-chemical properties and the structure of the genetic code. While conservation is the result of negative selection against proteins with reduced functionality, the codon sequences determine the stochastic aspect of amino acid exchanges. Thus far, it is known that the genetic code is the dominant factor if little time has elapsed since the divergence of one gene into two, but physico-chemical forces gain importance at greater evolutionary distances. Further details, however, on how the influence of these factors varies with time are unknown to date. METHODOLOGY/PRINCIPAL FINDINGS: Here, we derive each 10,000 divergence specific substitution matrices for orthologues and paralogues from the Pfam collection of multiple protein alignments and quantify the action of three physico-chemical forces and of the structure of the genetic code at high resolution using correlation analysis. For closely related proteins, the codon sequence similarity is the most influential factor controlling protein evolution, but its influence decreases rapidly as divergence grows. From a protein sequence divergence of about 20 percent on the maintenance of the hydrophobic character of an amino acid is the most influential factor. All factors lose importance from about 40 percent divergence on. This suggests that the original protein structure often does no longer represent a constraint to the protein sequence. The proteins then become free to adopt new functions. We furthermore show that the constraints exerted by both physico-chemical forces and by the genetic code are quite comparable for orthologues and paralogues, however somewhat weaker for paralogues than for orthologues in weakly or moderately diverged proteins. CONCLUSION/SIGNIFICANCE: Our analysis substantiates earlier findings that protein evolution is mainly governed by the structure of the genetic code in the early phase after divergence and by the conservation of physico-chemical properties at the later phase. We determine the level of sequence divergence from which on the conservation of the hydrophobic character is gaining importance over the genetic code to be 20 percent. The evolution of orthologues and paralogues is shaped by evolutionary forces in quite comparable ways.