The acceptor substrate specificity of human beta 4-galactosyltransferase Vindicates its potential function in O-glycosylation

In order to assess the function of the different human UDP-Gal:GlcNAc beta 4-galactosyltransferases, the cDNAs of two of them, beta 4-GalT I and beta 4-GalT V, were expressed in the baculovirus/insect cell expression system, The soluble recombinant enzymes produced mere purified from the medium and used to determine their in vitro substrate specificities. The specific acti vity of the recombinant beta 4-GalT V was more than 15 times lower than tha t of beta 4-GalT I, using GlcNAc beta-S-pNP as an acceptor. Whereas beta 4- GalT I efficiently acts on all substrates hating a terminal beta-linked Glc NAc, beta 4-GalT V appeared to be far more restricted in acceptor usage. be ta 4-GalT V acts with high preference on accepters that contain the GlcNAc beta 1-->6GalNAc structural element, as found in O-linked core 2-, 4- and 6 -based glycans, but not on substrates related to N-linked or blood group I- active oligosaccharides. These results suggest that beta 4-GalT V may funct ion in the synthesis of lacNAc units on O-linked chains, particularly in ti ssues which do not express beta 4-GalT I, such as brain. (C) 1999 Federatio n of European Biochemical Societies.