title: Complex-type N-linked oligosaccharides of gp120 from human immunodeficiency virus type 1 contain sulfated N-acetylglucosamine.
abstract: The major envelope glycoproteins gp120 and gp41 of human immunodeficiency virus type 1, the causative agent for human AIDS, contain numerous N-linked oligosaccharides. We report here our discovery that N-acetylglucosamine residues within the complex-type N-linked oligosaccharides of both gp120 and its precursor, gp160, are sulfated. When human Molt-3 cells persistently infected with human T-cell leukemia virus IIIB were metabolically radiolabeled with 35SO4, gp160, gp120, and to some extent gp41 were radiolabeled. The 35SO4-labeled oligosaccharides were quantitatively released by N-glycanase treatment and were bound by immobilized Ricinus communis agglutinin I, a lectin that binds to terminal beta-galactosyl residues. The kinetics of release of sulfate upon acid hydrolysis from 35SO4-labeled gp120 indicate that sulfation occurs in a primary sulfate ester linkage. Methylation analysis of total glycopeptides from Molt-3 cells metabolically radiolabeled with [3H]glucosamine demonstrates that sulfation occurs at the C-6 position of N-acetylglucosamine. Fragmentation of the gp120-derived 35SO4-labeled glycopeptides by treatment with hydrazine and nitrous acid and subsequent reduction generated galactosyl-anhydromannitol-6-35SO4, which is the expected reaction product from GlcNAc-6-sulfate within a sulfated lactosamine moiety. Charge analysis of the [3H]galactose- and [3H]glucosamine-labeled glycopeptides from gp120 and gp160 indicates that approximately 14% of the complex-type N-linked oligosaccharides are sulfated.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Oklahoma Health Science Center, Oklahoma City 73104.
authors.forename: A
authors.initials: A
authors.lastname: Shilatifard
authors.forename: R K
authors.initials: RK
authors.lastname: Merkle
authors.forename: D E
authors.initials: DE
authors.lastname: Helland
authors.forename: J L
authors.initials: JL
authors.lastname: Welles
authors.forename: W A
authors.initials: WA
authors.lastname: Haseltine
authors.forename: R D
authors.initials: RD
authors.lastname: Cummings
mesh_terms: D000117:Acetylglucosamine / Q000032:analysis*; D000581:Amidohydrolases / Q000378:metabolism; D002240:Carbohydrate Sequence; D002478:Cells, Cultured; D005690:Galactose / Q000378:metabolism; D015686:Gene Products, env / Q000378:metabolism; D006023:Glycoproteins / Q000737:chemistry; D015699:HIV Envelope Protein gp120 / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D019302:HIV Envelope Protein gp160; D015700:HIV Envelope Protein gp41 / Q000737:chemistry; D015497:HIV-1 / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D006834:Hydrazines / Q000494:pharmacology; D006868:Hydrolysis; D008745:Methylation; D008969:Molecular Sequence Data; D009608:Nitrous Acid / Q000494:pharmacology; D009844:Oligosaccharides / Q000096:biosynthesis / Q000737:chemistry*; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D011498:Protein Precursors / Q000378:metabolism; D012276:Ricin / Q000378:metabolism; D013431:Sulfates / Q000378:metabolism; D013464:Sulfuric Acids / Q000032:analysis*
chemical_list: D015686:Gene Products, env; D006023:Glycoproteins; D015699:HIV Envelope Protein gp120; D019302:HIV Envelope Protein gp160; D015700:HIV Envelope Protein gp41; D006834:Hydrazines; D009844:Oligosaccharides; D011498:Protein Precursors; D013431:Sulfates; D013464:Sulfuric Acids; C029424:hydrazine; D012276:Ricin; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D009608:Nitrous Acid; D000117:Acetylglucosamine; D005690:Galactose