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Biosynthesis of Glucosyl Glycerol,a Compatible Solute,Using Intermolecular Transglycosylation Activity of Amylosucrase from Methylobacillus flagellatus KT
Authors:Jin-Woo Jeong  Dong-Ho Seo  Jong-Hyun Jung  Ji-Hae Park  Nam-In Baek  Myo-Jeong Kim  Cheon-Seok Park
Institution:1. Graduate School of Biotechnology and Institute of Life Science and Resources, Kyung Hee University, Yongin, 446-701, South Korea
2. Food Research Institute and School of Food and Life Science and Biohealth Products Research Center, Inje University, Gimhae, 621-749, South Korea
Abstract:A putative α-amylase gene (accession number, CP000284) of Methylobacillus flagellatus KT ATCC51484 was cloned in Escherichia coli, and its gene product was expressed and characterized. The purified recombinant enzyme (MFAS) displayed a typical amylosucrase (ASase) activity by the demonstration of multiple activities of hydrolysis, isomerization, and polymerization although it was designated as an α-amylase. The optimal reaction temperature and pH for the sucrose hydrolysis activity of MFAS were determined to be 45 °C and pH 8.5, respectively. MFAS has relatively high thermostable characteristics compared with other ASases, as demonstrated by a half-life of 19.3 min at 50 °C. MFAS also showed polymerization activity using sucrose as a sole substrate. Glycerol was transglycosylated by the intermolecular transglycosylation activity of MFAS. Two major products were observed by thin-layer chromatography and isolated by paper chromatography and recycling HPLC. Using 1H and 13C NMR, their chemical structures were determined to be (2S)-1-O-α-d-glucosyl-glycerol or (2R)-1-O-α-d-glucosyl-glycerol and 2-O-α-d-glucosyl-glycerol, in which a glucose molecule is linked to glycerol via an α-glycosidic linkage.
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