Mucin-type O-glycosylation is controlled by short- and long-range glycopeptide substrate recognition that varies among members of the polypeptide GalNAc transferase family

Glycobiology. 2016 Apr;26(4):360-76. doi: 10.1093/glycob/cwv108. Epub 2015 Nov 26.

Abstract

A large family of UDP-GalNAc:polypeptide GalNAc transferases (ppGalNAc-Ts) initiates and defines sites of mucin-type Ser/Thr-O-GalNAc glycosylation. Family members have been classified into peptide- and glycopeptide-preferring subfamilies, although both families possess variable activities against glycopeptide substrates. All but one isoform contains a C-terminal carbohydrate-binding lectin domain whose roles in modulating glycopeptide specificity is just being understood. We have previously shown for several peptide-preferring isoforms that the presence of a remote Thr-O-GalNAc, 6-17 residues from a Ser/Thr acceptor site, may enhance overall catalytic activity in an N- or C-terminal direction. This enhancement varies with isoform and is attributed to Thr-O-GalNAc interactions at the lectin domain. We now report on the glycopeptide substrate utilization of a series of glycopeptide (human-ppGalNAc-T4, T7, T10, T12 and fly PGANT7) and peptide-preferring transferases (T2, T3 and T5) by exploiting a series of random glycopeptide substrates designed to probe the functions of their catalytic and lectin domains. Glycosylation was observed at the -3, -1 and +1 residues relative to a neighboring Thr-O-GalNAc, depending on isoform, which we attribute to specific Thr-O-GalNAc binding at the catalytic domain. Additionally, these glycopeptide-preferring isoforms show remote lectin domain-assisted Thr-O-GalNAc enhancements that vary from modest to none. We conclude that the glycopeptide specificity of the glycopeptide-preferring isoforms predominantly resides in their catalytic domain but may be further modulated by remote lectin domain interactions. These studies further demonstrate that both domains of the ppGalNAc-Ts have specialized and unique functions that work in concert to control and order mucin-type O-glycosylation.

Keywords: glycoprotein biosynthesis; glycosyltransferase; lectin; mucin.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence / genetics
  • Binding Sites
  • Carbohydrates / chemistry
  • Carbohydrates / genetics
  • Catalytic Domain
  • Fucose / analogs & derivatives
  • Fucose / chemistry
  • Glycopeptides / biosynthesis
  • Glycopeptides / chemistry*
  • Glycopeptides / genetics
  • Glycosylation
  • Humans
  • Lectins / chemistry*
  • Lectins / genetics
  • Mucins / biosynthesis
  • Mucins / chemistry*
  • Mucins / genetics
  • Phylogeny
  • Protein Binding
  • Protein Conformation
  • Protein Structure, Tertiary
  • Sialyltransferases / chemistry*
  • Sialyltransferases / genetics
  • Substrate Specificity

Substances

  • Carbohydrates
  • Glycopeptides
  • Lectins
  • Mucins
  • Fucose
  • N-acetylfucosamine
  • Sialyltransferases