Isoform-specific O-glycosylation by murine UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase-T3, in vivo.
Multiple isoforms of UDP-GalNAc:polypeptide N-acetylgalactosaminyl- transferase (ppGaNTase) have been cloned and expressed from a variety of organisms. In general, these isoforms display different patterns of tissue-specific expression, but exhibit overlapping substrate specificities, in vitro . A peptide substrate, derived from the sequence of the V3 loop of the HIV gp120 protein (HIV peptide), has previously been shown to be glycosylated in vitro exclusively by the ppGaNTase-T3 (Bennett et al. , 1996). To determine if this isoform-specificity is maintained in vivo , we have examined the glycosylation of this substrate when it is expressed as a reporter peptide (rHIV) in a cell background (COS7 cells) which lacks detectable levels of the ppGaNTase-T3. Glycosylation of rHIV was greatly increased by coexpression of a recombinant ppGaNTase-T3. Overexpression of ppGaNTase-T1 yielded only partial glycosylation of the reporter. We have also determined that the introduction of a proline residue at the +3 position flanking the potential glycosylation site eliminated ppGaNTase-T3 selectivity toward rHIV observed both in vivo and in vitro .
Nehrke
K
K
Department of Dental Research, School of Dentistry, University of Rochester, Rochester, NY 14648, USA.
Hagen
F K
FK
Tabak
L A
LA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D019556:COS Cells; D006031:Glycosylation; D015699:HIV Envelope Protein gp120 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D017350:N-Acetylgalactosaminyltransferases / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D000097763:Polypeptide N-acetylgalactosaminyltransferase
D015699:HIV Envelope Protein gp120; D007527:Isoenzymes; D017350:N-Acetylgalactosaminyltransferases; D010446:Peptide Fragments; D011994:Recombinant Proteins; D000097763:Polypeptide N-acetylgalactosaminyltransferase; C059762:HIV envelope protein gp120 (305-321)