title: UDP-sulfoquinovose formation by Sulfolobus acidocaldarius.
abstract: The UDP-sulfoquinovose synthase Agl3 from Sulfolobus acidocaldarius converts UDP-D-glucose and sulfite to UDP-sulfoquinovose, the activated form of sulfoquinovose required for its incorporation into glycoconjugates. Based on the amino acid sequence, Agl3 belongs to the short-chain dehydrogenase/reductase enzyme superfamily, together with SQD1 from Arabidopsis thaliana, the only UDP-sulfoquinovose synthase with known crystal structure. By comparison of sequence and structure of Agl3 and SQD1, putative catalytic amino acids of Agl3 were selected for mutational analysis. The obtained data suggest for Agl3 a modified dehydratase reaction mechanism. We propose that in vitro biosynthesis of UDP-sulfoquinovose occurs through an NAD(+)-dependent oxidation/dehydration/enolization/sulfite addition process. In the absence of a sulfur donor, UDP-D-glucose is converted via UDP-4-keto-D-glucose to UDP-D-glucose-5,6-ene, the structure of which was determined by (1)H and (13)C-NMR spectroscopy. During the redox reaction the cofactor remains tightly bound to Agl3 and participates in the reaction in a concentration-dependent manner. For the first time, the rapid initial electron transfer between UDP-D-glucose and NAD(+) could be monitored in a UDP-sulfoquinovose synthase. Deuterium labeling confirmed that dehydration of UDP-D-glucose occurs only from the enol form of UDP-4-keto-glucose. The obtained functional data are compared with those from other UDP-sulfoquinovose synthases. A divergent evolution of Agl3 from S. acidocaldarius is suggested.
authors.affiliation: Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Muthgasse 11, 1190, Vienna, Austria.
authors.forename: Behnam
authors.initials: B
authors.lastname: Zolghadr
authors.forename: Bernhard
authors.initials: B
authors.lastname: Gasselhuber
authors.forename: Markus
authors.initials: M
authors.lastname: Windwarder
authors.forename: Martin
authors.initials: M
authors.lastname: Pabst
authors.forename: Daniel
authors.initials: D
authors.lastname: Kracher
authors.forename: Martina
authors.initials: M
authors.lastname: Kerndl
authors.forename: Sonja
authors.initials: S
authors.lastname: Zayni
authors.forename: Andreas
authors.initials: A
authors.lastname: Hofinger-Horvath
authors.forename: Roland
authors.initials: R
authors.lastname: Ludwig
authors.forename: Dietmar
authors.initials: D
authors.lastname: Haltrich
authors.forename: Chris
authors.initials: C
authors.lastname: Oostenbrink
authors.forename: Christian
authors.initials: C
authors.lastname: Obinger
authors.forename: Paul
authors.initials: P
authors.lastname: Kosma
authors.forename: Paul
authors.initials: P
authors.lastname: Messner
authors.forename: Christina
authors.initials: C
authors.lastname: Schäffer
mesh_terms: D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005964:Glucosyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009243:NAD / Q000378:metabolism; D016944:Sulfolobus / Q000378:metabolism*; D014532:Uridine Diphosphate Glucose / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism
chemical_list: D019843:Archaeal Proteins; C096386:uridine diphosphate sulfoquinovose; D009243:NAD; D005964:Glucosyltransferases; D014532:Uridine Diphosphate Glucose