Variability‐based sequence alignment identifies residues responsible for functional differences in α and β tubulin

Abstract

α and β Tubulin are well‐characterized paralogs with similar structures and functions. We quantify the variability of every amino acid position in both tubulins from the aligned sequences of their numerous known orthologs. By aligning the variability profiles, we identify residues that differ significantly in variability between α and β tubulin. Most of these residues are part of well‐defined secondary structures and are clustered around the nucleotide binding pocket, the site of greatest functional difference between the two paralogs. The remaining residues of large difference in variability are located in the N‐terminal loop between H1 and S2. We therefore predict that certain residues in this unstructured region also contribute to a functional difference between α and β tubulin. Furthermore, we find the most restrictive variability‐based alignment is nearly identical to the true structure‐based alignment. Thus, by using a stringent variability‐based alignment to approximate the true alignment, the method introduced here may predict sites of functional distinction between paralogous proteins even in the absence of structural information.