Structural comparisons of TIM barrel proteins suggest functional and evolutionary relationships between beta-galactosidase and other glycohydrolases

beta-Galactosidase (lacZ) from Escherichia coli is a 464 kDa homotetramer. Each subunit consists of five domains, the third being an alpha/beta barrel that contains most of the active site residues. A comparison is made betwe en each of the domains and a large set of proteins representative of all st ructures from the protein data bank. Many structures include an alpha/beta barrel. Those that are most similar to the alpha/beta barrel of E. coli bet a-galactosidase have similar catalytic residues and belong to the so-called "4/7 superfamily" of glycosyl hydrolases. The structure comparison suggest s that beta-amylase should also be included in this family. Of three struct ure comparison methods tested, the "ProSup" procedure of Zu-Kang and Sippl and the "Superimpose" procedure of Diederichs were slightly superior in dis criminating the members of this superfamily, although all procedures were v ery powerful in identifying related protein structures. Domains 1, 2, and 4 of E. coli beta-galactosidase have topologies related to "jelly-roll barre ls" and "immunoglobulin constant" domains. This fold also occurs in the cel lulose binding domains (CBDs) of a number of glycosyl hydrolases. The fold of domain 1 of E. coli beta-galactosidase is closely related to some CBDs, and the domain contributes to substrate binding, but in a manner unrelated to cellulose binding by the CBDs. This is typical of domains 1, 2, 4, and 5 , which appear to have been recruited to play roles in beta-galactosidase t hat are unrelated to the functions that such domains provide in other conte xts. It is proposed that beta-galactosidase arose from a prototypical singl e domain alpha/beta barrel with an extended active site cleft. The subseque nt incorporation of elements from other domains could then have reduced the size of the active site from a cleft to a pocket to better hydrolyze the d isaccharide lactose and, at the same time, to facilitate the production of inducer, allolactose.