Ancestral protein reconstruction: techniques and applications.

Ancestral sequence reconstruction (ASR) is the calculation of ancient protein sequences on the basis of extant ones. It is most powerful in combination with the experimental characterization of the corresponding proteins. Such analyses allow for the study of problems that are otherwise intractable. For example, ASR has been used to characterize ancestral enzymes dating back to the Paleoarchean era and to deduce properties of the corresponding habitats. In addition, the historical approach underlying ASR enables the identification of amino acid residues key to protein function, which is often not possible by only comparing extant proteins. Along these lines, residues responsible for the spectroscopic properties of protein pigments were identified as well as residues determining the binding specificity of steroid receptors. Further applications are studies related to the longevity of mutations, the contribution of gene duplications to enzyme functionalization, and the evolution of protein complexes. For these applications of ASR, we discuss recent examples; moreover, we introduce the basic principles of the underlying algorithms and present state-of-the-art protocols.