Cloning of Gb sub(3) Synthase, the Key Enzyme in Globo-series Glycosphingolipid Synthesis, Predicts a Family of alpha 1,4-Glycosyltransferases Conserved in Plants, Insects, and Mammals

• Published in: The Journal of biological chemistry 2000-08, Vol.275 (33), p.25315-25321
• Main Authors: Keusch, J J, Manzella, S M, Nyame, KA, Cummings, R D, Baenziger, JU
• Format: Article
• Language: English
• Subjects: a1,4-glactosyltransferase; Arabidopsis; Drosophila melanogaster; Gb3 synthase; glycosphingolipids; verotoxin
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M002630200

We have cloned Gb sub(3) synthase, the key alpha 1,4-galactosyltransferase in globo-series glycosphingolipid (GSL) synthesis, via a phenotypic screen, which previously yielded iGb sub(3) synthase, the alpha 1,3-galactosyltransferase required in isoglobo-series GSL (Keusch, J. J., Manzella, S. M., Nyame, K. A., Cummings, R. D., and Baenziger, J. U. (2000) J. Biol. Chem. 33). Both transferases act on lactosylceramide, Gal beta 1,4Glc beta 1Cer (LacCer), to produce Gb sub(3) (Gal alpha 1,4LacCer) or iGb sub(3) (Gal alpha 1,3LacCer), respectively. GalNAc can be added sequentially to either Gb sub(3) or iGb sub(3) yielding globoside and Forssman from Gb sub(3), and isogloboside and isoForssman from iGb sub(3). Gb sub(3) synthase is not homologous to iGb sub(3) synthase but shows 43% identity to a human alpha 1,4GlcNAc transferase that transfers a UDP-sugar in an alpha 1,4-linkage to a beta -linked Gal found in mucin. Extensive homology (35% identity) is also present between Gb sub(3) synthase and genes in Drosophila melanogaster and Arabidopsis thaliana, supporting conserved expression of an alpha 1,4-glycosyltransferase, possibly Gb sub(3) synthase, throughout evolution. The isolated Gb sub(3) synthase cDNA encodes a type II transmembrane glycosyltransferase of 360 amino acids. The highest tissue expression of Gb sub(3) synthase RNA is found in the kidney, mesenteric lymph node, spleen, and brain. Gb sub(3) glycolipid, also called P antigen or CD77, is a known receptor for verotoxins. CHO cells that do not express Gb sub(3) and are resistant to verotoxin become susceptible to the toxin following transfection with Gb sub(3) synthase cDNA.