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 (Gal1-4GlcNAc) but not lacto-N-biose (Gal1-3GlcNAc) 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 (Gal1-4GlcNAc) contribute to the interaction with GlcNAc moiety. These three residues play a key role in establishing the acceptor substrate specificity.
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