title: Synthesis of a fluorescent acceptor substrate for glycosyltransferases involved in the assembly of O-antigens of enterohemorrhagic Escherichia coli O157 and O5.
abstract: The assembly of the repeating units of O-antigens in Gram negative bacteria is catalyzed by specific glycosyltransferases. Previously we used GlcNAc/GalNAcα-diphosphate-phenoxyundecyl as natural acceptor substrate analogs in assays of the transfer of radioactive sugars by bacterial glycosyltransferases. In order to develop new, fluorescence based assays we have synthesized a fluorescent acceptor P¹-[11-(anthracen-9-ylmethoxy)undecyl]-P²-(2-acetamido-2-deoxy-α-D-galactopyranosyl) diphosphate and have shown that the compound was an excellent acceptor for glucosyltransferase WbdN from Escherichia coli (E. coli) O157 and for galactosyltransferase WbwC from E. coli O5. This is the first report of the Gal-transferase activity of the wbwC gene product of E.coli O5. The presence of the fluorescent label in the acceptor molecule allows the detection of glycosyltransferase reaction products with high sensitivity, eliminating the need for radioactive nucleotide sugars.
authors.affiliation: N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, Moscow, Russia.
authors.forename: Anna N
authors.initials: AN
authors.lastname: Vinnikova
authors.forename: Tatyana N
authors.initials: TN
authors.lastname: Druzhinina
authors.forename: Leonid L
authors.initials: LL
authors.lastname: Danilov
authors.forename: Natalia S
authors.initials: NS
authors.lastname: Utkina
authors.forename: Vladimir I
authors.initials: VI
authors.lastname: Torgov
authors.forename: Vladimir V
authors.initials: VV
authors.lastname: Veselovsky
authors.forename: Shuo
authors.initials: S
authors.lastname: Wang
authors.forename: Bin
authors.initials: B
authors.lastname: Liu
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
authors.forename: Inka
authors.initials: I
authors.lastname: Brockhausen
mesh_terms: D055162:Biocatalysis; D054324:Enterohemorrhagic Escherichia coli / Q000201:enzymology / Q000378:metabolism*; D019453:Escherichia coli O157 / Q000201:enzymology / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism*; D016695:Glycosyltransferases / Q000378:metabolism*; D015394:Molecular Structure; D019081:O Antigens / Q000096:biosynthesis* / Q000737:chemistry*; D013237:Stereoisomerism
chemical_list: D005456:Fluorescent Dyes; D019081:O Antigens; D016695:Glycosyltransferases