title: The effect of heavy atoms on the conformation of the active-site polypeptide loop in human ABO(H) blood-group glycosyltransferase B.
abstract: The human ABO(H) blood-group antigens are oligosaccharide structures that are expressed on erythrocyte and other cell surfaces. The terminal carbohydrate residue differs between the blood types and determines the immune reactivity of this antigen. Individuals with blood type A have a terminal N-acetylgalactosamine residue and those with blood type B have a terminal galactose residue. The attachment of these terminal carbohydrates are catalyzed by two different glycosyltransferases: an alpha(1-->3)N-acetylgalactosaminyltransferase (GTA) and an alpha(1-->3)galactosyltransferase (GTB) for blood types A and B, respectively. GTA and GTB are homologous enzymes that differ in only four of 354 amino-acid residues (Arg/Gly176, Gly/Ser235, Leu/Met266 and Gly/Ala268 in GTA and GTB, respectively). Diffraction-quality crystals of GTA and GTB have previously been grown from as little as 10 mg ml(-1) stock solutions in the presence of Hg, while diffraction-quality crystals of the native enzymes require much higher concentrations of protein. The structure of a single mutant C209A has been determined in the presence and absence of heavy atoms and reveals that when mercury is complexed with Cys209 it forces a significant level of disorder in a polypeptide loop (amino acids 179-195) that is known to cover the active site of the enzyme. The observation that the more highly disordered structure is more amenable to crystallization is surprising and the derivative provides insight into the mobility of this polypeptide loop compared with homologous enzymes.
authors.affiliation: The Department of Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, Canada.
authors.forename: James A
authors.initials: JA
authors.lastname: Letts
authors.forename: Mattias
authors.initials: M
authors.lastname: Persson
authors.forename: Brock
authors.initials: B
authors.lastname: Schuman
authors.forename: Svetlana N
authors.initials: SN
authors.lastname: Borisova
authors.forename: Monica M
authors.initials: MM
authors.lastname: Palcic
authors.forename: Stephen V
authors.initials: SV
authors.lastname: Evans
mesh_terms: D000017:ABO Blood-Group System / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D005700:Galactosyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D040681:Structural Homology, Protein
chemical_list: D000017:ABO Blood-Group System; D010455:Peptides; D005700:Galactosyltransferases; C023155:blood-group-substance alpha-D-galactosyltransferase; D003545:Cysteine