Real-time NMR monitoring of intermediates and labile products of the bifunctional enzyme UDP-apiose/UDP-xylose synthase.
The conversion of UDP-alpha-d-glucuronic acid to UDP-alpha-d-xylose and UDP-alpha-d-apiose by a bifunctional potato enzyme UDP-apiose/UDP-xylose synthase was studied using real-time nuclear magnetic resonance (NMR) spectroscopy. UDP-alpha-d-glucuronic acid is converted via the intermediate uridine 5'-beta-l-threo-pentapyranosyl-4''-ulose diphosphate to UDP-alpha-d-apiose and simultaneously to UDP-alpha-d-xylose. The UDP-alpha-d-apiose that is formed is unstable and is converted to alpha-d-apio-furanosyl-1,2-cyclic phosphate and UMP. High-resolution real-time NMR spectroscopy is a powerful tool for the direct and quantitative characterization of previously undetected transient and labile components formed during a complex enzyme-catalyzed reaction.
Complex Carbohydrate Research Center, and BioEnergy Science Center, 315 Riverbend Road, University of Georgia, Athens, GA 30602, USA.
Paul
P
Guyett
John
J
Glushka
Xiaogang
X
Gu
Maor
M
Bar-Peled
D055162:Biocatalysis; D002262:Carboxy-Lyases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011198:Solanum tuberosum / Q000201:enzymology; D013997:Time Factors; D014535:Uridine Diphosphate Glucuronic Acid / Q000378:metabolism; D014539:Uridine Diphosphate Sugars / Q000378:metabolism; D014994:Xylose / Q000031:analogs & derivatives / Q000378:metabolism
D011994:Recombinant Proteins; C511616:UDP-apiose; D014539:Uridine Diphosphate Sugars; D014535:Uridine Diphosphate Glucuronic Acid; D014994:Xylose; D002262:Carboxy-Lyases; C023257:UDPglucuronate decarboxylase