Protein Science zyxwvutsrqpon (1994), 3:2045-2054. Cambridge University Press. Printed zyxwvuts in the USA Copyright zyxwvutsrqp 0 1994 The Protein Society Eukaryotic translation elongation factor ly contains a glutathione transferase domain zyx - Study of a diverse, ancient protein superfamily using motif search and structural modeling EUGENE V. KOONIN,’ ARCADY R. MUSHEGIAN,2*4 ROMAN L. TATUSOV,’ STEPHEN F. ALTSCHUL,’ STEPHEN H. BRYANT,’ PEER BORK,3 AND ALFONSO VALENCIA3 ’ National Center for Biotechnology Information, National Library of Medicine, * Department of Plant Pathology, University of Kentucky, Lexington, Kentucky 40546-0091 National Institutes of Health, Bethesda, Maryland 20894 European Molecular Biology Laboratory, Meyerhofstrasse zyxwvuts 1, D-6900, Heidelberg, Germany (RECEIVED June 23, 1994; ACCEPTED August 9, 1994) Abstract Using computer methods for multiple alignment, sequence motif search, and tertiary structuremodeling, we show that eukaryotic translation elongation factor ly (EFly) contains an N-terminal domain related to class 0 gluta- thione S-transferases (GST). GST-like proteins related to class 8 comprise a large group including, in addition to typical GSTs and EFly, stress-induced proteins from bacteria and plants, bacterial reductive dehalogenases and P-etherases, and several uncharacterized proteins. These proteins share 2 conserved sequence motifs with GSTs of other classes zyxwvutsr (a, zyxwvutsrq p, and K). Tertiary structure modeling showed that in spite of the relatively low sequence sim- ilarity, the GST-related domain of EFly is likely to form a fold very similar to that in the known structures of class zyxwvutsrqpon a, p, and ?r GSTs. One of the conserved motifs is implicated in glutathione binding, whereas the other mo- tif probably is involved in maintaining the proper conformation of the GST domain. We predict that the GST- like domain in EFly is enzymatically active and that to exhibit GST activity, EFly has to form homodimers. The GST activity may be involved in the regulation of the assembly of multisubunit complexes containing EFl and aminoacyl-tRNA synthetases by shifting the balance between glutathione, disulfide glutathione, thiol groups of cysteines, and protein disulfide bonds. The GST domain is a widespread, conserved enzymatic module that may be covalently or noncovalently complexed with other proteins. Regulation of protein assembly and folding may be 1 of the functions of GST. Keywords: conserved sequence motifs; glutathione S-transferase domain; motif search; structure modeling; trans- lation elongation factor ly Combination of functionally distinct domains in a single poly- peptide is one of the general principles in the build-up of com- plex biochemical systems (Bork, 1992; Doolittle, 1992; Doolittle & Bork, 1993). In particular, several widespread enzymatic do- mains are known that may be combined with a variety of other domains and provide a function that is common to different pro- cesses. Examples of such universal domains include ATPase Reprint requests to: Eugene V. Koonin, National Center for Biotech- nology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894; e-mail: koonin@ncbi.nlm. nih.gov. Present address: Department of Microbiology, University of Wash- ington, Seattle, Washington 98195. (Gorbalenya & Koonin, 1990; Milner-White et al., 1991), pro- tein kinase (Hanks et al., 1988), and serine protease (Neurath, 1986). We show here that glutathione S-transferase (GST) may be another “portable”enzymatic domain. GSTs are dimeric pro- teins that catalyze the conjugation of glutathione (GSH) with a variety of electrophiles, according to the equation GSH + EN = GS-E + NH, where EN is an electrophilic substrate (re- viewed in Pickett & Lu, 1989; Fahey & Sundquist, 1991; Pem- ble & Taylor, 1992; Rushmore & Pickett, 1993; Dirr et al., 1994; Wilce & Parker, 1994). The best studied electrophilic substrates include chlorinated compounds and other xenobiotics, epoxides, and peroxides, e.g., hydrogen peroxide. It has been suggested that protection of cells against oxygen toxicity may be the pri- 2045