The beta 1,3-galactosyltransferase beta 3GalT-V is a stage-specific embryonic antigen-3 (SSEA-3) synthase

We have previously reported the molecular cloning of beta 1,3-galactosyltra nsferase-V (P3GalT-V), which catalyzes the transfer of Gal to GlcNAc-based accepters with a preference for the core3 O-linked glycan GlcNAc(beta 1,3)- GalNAc structure. Further characterization indicated that the recombinant b eta 3GalT-V enzyme expressed in Sf9 insect cells also utilized the glycolip id Lc3Cer as an efficient acceptor. Surprisingly, we also found that beta 3 GalT-V catalyzes the transfer of Gal to the terminal GalNAc unit of the glo boside Gb4, thereby synthesizing the glycolipid Gb5, also known as the stag e-specific embryonic antigen-3 (SSEA-3). The SSEA-3 synthase activity of be ta 3GalT-V was confirmed in vivo by stable expression of the human beta 3Ga lT-V gene in F9 mouse teratocarcinoma cells, as detected with the monoclona l antibody MC-631 by flow cytometry analysis and immunostaining of extracte d glycolipids. The biological relation between SSEA-3 formation and beta 3G alT-V was further documented by showing that F9 cells treated with the diff erentiation-inducing agent retinoic acid induced the expression of both the SSEA-3 epitope and the endogenous mouse beta 3GalT-V gene. This study repr esents the first example of a glycosyltransferase, which utilizes two kinds of sugar acceptor substrates without requiring any additional modifier mol ecule.