Cloning of Gb(3) synthase, the key enzyme in globo-series glycosphingolipid synthesis, predicts a family of alpha 1,4-glycosyltransferases conserved in plants, insects, and mammals

We have cloned Gb, synthase, the key alpha 1,4-galactosyl-transferase in gl obe-series glycosphingolipid (GSL) synthesis, via a phenotypic screen, whic h previously yielded iGb(3) synthase, the alpha 1,3-galactosyltransferase r equired in iso-globe-series GSL (Keusch, J,J., Manzella, S. M., Nyame, K. A ., Cummings, R. D., and Baenziger, J. U. (2000) J. Biol. Chem. 33), Both tr ansferases act on lactosylceramide, Gal beta 1,4Glc beta 1Cer (LacCer), to produce Gb(3) (Gal alpha 1,4LacCer) or iGb(3) (Ga1 alpha 1,3LacCer), respec tively. GalNAc can be added sequentially to either Gb(3) or iGb(3) yielding globoside and Forssman from Gb,, and isogloboside and isoForssman hem iGb( 3). Gb, synthase is not homologous to iGb(3) synthase but shows 43% identit y to a human alpha 1,4GlcNAc transferase that transfers a UDP-sugar in an a lpha 1,4-linkage to a beta-linked Gal found in mucin. Extensive homology (3 5% identity) is also present between Gb, synthase and genes in Drosophila m elanogaster and Arabidopsis thaliana, supporting conserved expression of an alpha 1,4-glycosyltransferase, possibly Gb, synthase, throughout evolution . The isolated Gb, synthase cDNA encodes a type II transmembrane glycosyltr ansferase of 360 amino acids. The highest tissue expression of Gb, synthase RNA is found in the kidney, mesenteric lymph node, spleen, and brain. Gb, glycolipid, also called pk antigen or CD77, is a known receptor for verotox ins. CHO cells that do not express Gb, and are resistant to verotoxin becom e susceptible to the toxin following transfection with Gb, synthase cDNA.