Structural basis for acceptor substrate recognition of a human glucuronyltransferase, GlcAT-P, an enzyme critical in the biosynthesis of the carbohydrate epitope HNK-1

J Biol Chem. 2004 May 21;279(21):22693-703. doi: 10.1074/jbc.M400622200. Epub 2004 Mar 1.


The HNK-1 carbohydrate epitope is found on many neural cell adhesion molecules. Its structure is characterized by a terminal sulfated glucuronyl acid. The glucuronyltransferases, GlcAT-P and GlcAT-S, are involved in the biosynthesis of the HNK-1 epitope, GlcAT-P as the major enzyme. We overexpressed and purified the recombinant human GlcAT-P from Escherichia coli. Analysis of its enzymatic activity showed that it catalyzed the transfer reaction for N-acetyllactosamine (Galbeta1-4GlcNAc) but not lacto-N-biose (Galbeta1-3GlcNAc) as an acceptor substrate. Subsequently, we determined the first x-ray crystal structures of human GlcAT-P, in the absence and presence of a donor substrate product UDP, catalytic Mn(2+), and an acceptor substrate analogue N-acetyllactosamine (Galbeta1-4GlcNAc) or an asparagine-linked biantennary nonasaccharide. The asymmetric unit contains two independent molecules. Each molecule is an alpha/beta protein with two regions that constitute the donor and acceptor substrate binding sites. The UDP moiety of donor nucleotide sugar is recognized by conserved amino acid residues including a DXD motif (Asp(195)-Asp(196)-Asp(197)). Other conserved amino acid residues interact with the terminal galactose moiety of the acceptor substrate. In addition, Val(320) and Asn(321), which are located on the C-terminal long loop from a neighboring molecule, and Phe(245) contribute to the interaction with GlcNAc moiety. These three residues play a key role in establishing the acceptor substrate specificity.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Amino Acid Sequence
  • Amino Sugars / metabolism
  • Binding Sites
  • CD57 Antigens / biosynthesis*
  • CD57 Antigens / chemistry
  • Carbohydrates / chemistry
  • Catalysis
  • Cell Adhesion
  • Crystallography, X-Ray
  • Dimerization
  • Electrons
  • Epitopes
  • Escherichia coli / metabolism
  • Glucuronosyltransferase / biosynthesis
  • Glucuronosyltransferase / chemistry*
  • Humans
  • Manganese / metabolism
  • Models, Molecular
  • Molecular Sequence Data
  • Protein Binding
  • Protein Conformation
  • Protein Structure, Quaternary
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Substrate Specificity
  • Uridine Diphosphate / chemistry


  • Amino Sugars
  • CD57 Antigens
  • Carbohydrates
  • Epitopes
  • N-acetyllactosamine
  • Manganese
  • Uridine Diphosphate
  • galactosylgalactoylxylosylprotein 3-beta-glucuronosyltransferase
  • Glucuronosyltransferase

Associated data

  • PDB/1V82
  • PDB/1V83
  • PDB/1V84