The acceptor substrate specificity of human β4‐galactosyltransferase V indicates its potential function in O‐glycosylation

In order to assess the function of the different human UDP‐Gal:GlcNAc β4‐galactosyltransferases, the cDNAs of two of them, β4‐GalT I and β4‐GalT V, were expressed in the baculovirus/insect cell expression system. The soluble recombinant enzymes produced were purified from the medium and used to determine their in vitro substrate specificities. The specific activity of the recombinant β4‐GalT V was more than 15 times lower than that of β4‐GalT I, using GlcNAcβ‐S‐pNP as an acceptor. Whereas β4‐GalT I efficiently acts on all substrates having a terminal β‐linked GlcNAc, β4‐GalT V appeared to be far more restricted in acceptor usage. β4‐GalT V acts with high preference on acceptors that contain the GlcNAcβ1→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 β4‐GalT V may function in the synthesis of lacNAc units on O‐linked chains, particularly in tissues which do not express β4‐GalT I, such as brain.