ISOFORM-SPECIFIC O-GLYCOSYLATION BY MURINE UDP-GALNAC, POLYPEPTIDE N-ACETYLGALACTOSAMINYLTRANSFERASE-T3, IN-VIVO

Multiple isoforms of UDP-GalNAc:polypeptide N-acetylgalactosaminyl-tra nsferase (ppGaNTase) have been cloned and expressed from a variety of organisms, In general, these isoforms display different patterns of ti ssue-specific expression, but exhibit overlapping substrate specificit ies, in vitro, A peptide substrate, derived from the sequence of the V 3 loop of the HIV gp120 protein (HIV peptide), has previously been sho wn to be glycosylated in vitro exclusively by the ppGaNTase-T3 (Bennet t ef al., 1996), To determine if this isoform-specificity is maintaine d in vivo, we have examined the glycosylation of this substrate when i t is expressed as a reporter peptide (rHIV) in a cell background (COS7 cells) which lacks detectable levels of the ppGaNTase-T3, Glycosylati on of rHIV was greatly increased by coexpression of a recombinant ppGa NTase-T3, Overexpression of ppGaNTase-T1 yielded only partial glycosyl ation of the reporter, We have also determined that the introduction o f a proline residue at the +3 position flanking the potential glycosyl ation site eliminated ppGaNTase-T3 selectivity toward rHIV observed bo th in vivo and in vitro.