It has been argued that the limited set of proteins used by life as we know could not have arisen by the process of Darwinian selection from all possible proteins. This probabilistic argument has a number of implicit assumptions that may not be warranted. A variety of considerations are presented to show that the number of amino acid sequences that need to have been sampled during the evolution of proteins is far smaller than assumed by the argument.

The evolutionary history of leeches is employed as a general framework for understanding more than merely the systematics of this charismatic group of annelid worms, and serves as a basis for understanding blood-feeding related correlates ranging from the specifics of gut-associated bacterial symbionts to salivary anticoagulant peptides. A variety of medicinal leech families were examined for intraluminal crop bacterial symbionts. Species of Aeromonas and Bacteroidetes were characterized with DNA gyrase B and 16S rDNA. Bacteroidetes isolates were found to be much more phylogenetically diverse and suggested stronger evidence of phylogenetic correlation than the gammaproteobacteria. Patterns that look like co-speciation with limited taxon sampling do not in the full context of phylogeny. Bioactive compounds that are expressed as gene products, like those in leech salivary glands, have ‘passed the test’ of evolutionary selection. We produced and bioinformatically mined salivary gland EST libraries across medicinal leech lineages to experimentally and statistically evaluate whether evolutionary selection on peptides can identify structure-function activities of known therapeutically relevant bioactive compounds like antithrombin, hirudin and antistasin. The combined information content of a well corroborated leech phylogeny and broad taxonomic coverage of expressed proteins leads to a rich understanding of evolution and function in leech history.

Biotin and lipoic acid moieties are the covalently attached coenzyme cofactors of several multicomponent enzyme complexes that catalyze key metabolic reactions. Attachment of these moieties to the biotinyl- and lipoyl-dependent enzymes is post-translationally catalyzed by specific biotinylating and lipoylating protein enzymes. In Escherichia coli, two different enzymes, LplA and LipB, catalyze independent pathways for the lipoylation of the relevant enzymes, whereas only one enzyme, the BirA protein, is responsible for all the biotinylation. Counterparts of the E. coli BirA, LplA, and LipB enzymes have been previously identified in many organisms, but homology among the three families has never been reported. Computational analysis based on PSI-BLAST profiles and secondary structure predictions indicates, however, that lipoylating and biotinylating enzymes are evolutionarily related protein families containing a homologous catalytic module. Sequence conservation among the three families is very poor, but a single lysine residue is strictly conserved in all of them, which, according to the available X-ray crystal structure of the E. coli BirA protein, is expected to contribute to the binding of lipoic acid in the LplA and LipB enzymes.

Lipidation catalyzed by protein prenyltransferases is essential for the biological function of a number of eukaryotic proteins, many of which are involved in signal transduction and vesicular traffic regulation. Sequence similarity searches reveal that the α-subunit of protein prenyltransferases (PTα) is a member of the tetratricopeptide repeat (TPR) superfamily. This finding makes the three-dimensional structure of the rat protein farnesyltransferase the first structural model of a TPR protein interacting with its protein partner. Structural comparison of the two TPR domains in protein farnesyltransferase and protein phosphatase 5 indicates that variation in TPR consensus residues may affect protein binding specificity through altering the overall shape of the TPR superhelix. A general approach to evolutionary analysis of proteins with repetitive sequence motifs has been developed and applied to the protein prenyltransferases and other TPR proteins. The results suggest that all members in PTα family originated from a common multirepeat ancestor, while the common ancestor of PTα and other members of TPR superfamily is likely to be a single repeat protein.