Differential Recognition of the Type I and II H Antigen Acceptors by the Human ABO(H) Blood Group A and B Glycosyltransferases

Abstract

The human ABO(H) blood group A and B antigens are generated by the homologous glycosyltransferases A (GTA) and B (GTB), which add the monosaccharides GalNAc and Gal, respectively, to the cell-surface H antigens. In the first comprehensive structural Gly/Ala-268). As these enzymes both utilize the H antigen acceptors, the four critical residues had been thought to be involved strictly in donor recognition; however, we now report that acceptor binding and subsequent transfer are significantly influenced by two of these residues: Gly/Ser-235 and Leu/Met-266. Furthermore, these structures show that acceptor recognition is dominated by the central Gal residue despite the fact that the L-Fuc residue is required for efficient catalysis and give direct insight into the design of model inhibitors for GTA and GTB.

Structural Basis for the Recognition of Blood Group Trisaccharides by Norovirus

“…The findings of previous nuclear magnetic resonance and crystallographic studies suggest that the conformation of the B trisaccharide is more rigid in general (17). Interestingly, in our case, the conformations of both ␣-Fuc and ␤-Gal of the B trisaccharide were well defined in the electron density map and were very similar to their counterparts in the H trisaccharide in a previously described complex with N-acetylgalactosaminyl transferase (14).…”

High Resolution Structures of the Human ABO(H) Blood Group Enzymes in Complex with Donor Analogs Reveal That the Enzymes Utilize Multiple Donor Conformations to Bind Substrates in a Stepwise Manner

“…These enzymes are essential for normal cell development, and aberrant GT function can result in a number of infections and inflammatory disease states (1). Despite their functional and physiological diversity, GTs exhibit structural phenotype conservation even in the absence of sequence homology (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12). There are currently 97 GT families based on sequence identity (2,13), with almost all of the enzymes falling into two major fold types: GT-A and GT-B.…”

ABO(H) Blood Group A and B Glycosyltransferases Recognize Substrate via Specific Conformational Changes

“…Crystallization-All proteins were crystallized using conditions different from those reported previously (10,19,33,35,38,39). Whereas the first crystals of GTB were grown from relatively low protein concentrations (ϳ8 -15 mg/ml) and as a mercury derivative, the crystals in this paper were initially generated from higher protein concentrations (ϳ60 -75 mg/ml).…”

“…A nomenclature based on these four critical amino acid residues has been developed to describe GTA and GTB chimera, where GTA can be referred to as AAAA and GTB as BBBB with each letter corresponding to one critical residue in increasing order, such that the ABBB chimera would correspond to the GTB/G176R mutant enzyme and AABB would correspond to the GTB/G176R/S235G mutant enzyme. Critical residues Leu/Met 266 and Gly/Ala 268 have been shown to be responsible for discrimination between the two donor molecules (30 -32), whereas Gly/Ser 235 and Leu/Met 266 significantly impact acceptor recognition (33); however, the function of the conserved mutation Arg/Gly 176 has been elusive. Structural studies in the past have been hampered by the fact that Arg/Gly 176 lies at the edge of the internal disordered loop from residues 176 -195; however, the development of crystallization conditions for BBBB (GTB), ABBB, and AABB in the absence of heavy atoms permits a structural investigation of the influence of residue 176 on loop ordering and substrate binding.…”

“…When the original structure of GTB was solved in complex with the H antigen disaccharide, it was observed that the ␣-L-Fucp only provided a single contact to the enzyme (a hydrogen bond between the O-4-hydroxyl and the side chain of amino acid residue Asp 326 ) (19). Structures of GTA and GTB reported in complex with seven different fragments and analogs of the H antigen acceptors revealed many novel aspects of acceptor recognition by these homologous enzymes (33); however, the absolute necessity of the ␣-L-Fucp for catalytic activity was obscure.…”