title: [Effects of two UDP-glucose dehydrogenases on hyaluronic acid biotransformation].
abstract: We amplified genes encoding UDP-glucose dehydrogenase, ecohasB from Escherichia coli and spyhasB from Streptococcus pyogenes. Both ecohasB and spyhasB were inserted into T7 expression vector pRX2 to construct recombinant plasmids pRXEB and pRXSB, and to express in E. coli BL21(DE3). After nickel column purification of UDP-glucose dehydrogenases, the enzymes were characterized. The optimum reaction condition of spyHasB was at 30 °C and pH 10. The specific activity reached 12.2 U/mg under optimum condition. The optimum reaction condition of ecoHasB was at 30 °C and pH 9. Its specific activity reached 5.55 U/mg under optimum condition. The pmuhasA gene encoding hyaluronic acid synthase was amplified from Pasteurella multocida and ligated with ecohasB and spyhasB to construct the coexpression vectors pBPAEB and pBPASB, respectively. The co-expression vectors were transformed into E. coli BW25113. Hyaluronic acid (HA) was produced by biotransformation and the conditions were optimized. When recombinant strains were used to produce hyaluronic acid, the higher the activity of UDP-glucose dehydrogenase was, the better its stability was, and the higher the HA production could reach. Under the optimal conditions, the yields of HA produced by pBPAEB/BW25113 and pBPASB/BW25113 in shake flasks were 1.52 and 1.70 g/L, respectively, and the production increased more than 2-3 folds as previously reported.
authors.forename: Donghui
authors.initials: D
authors.lastname: GuoI
authors.forename: Jian
authors.initials: J
authors.lastname: Han
authors.forename: Weifeng
authors.initials: W
authors.lastname: Liu
authors.forename: Zhenzhou
authors.initials: Z
authors.lastname: Fu
authors.forename: Qizhong
authors.initials: Q
authors.lastname: Zhu
authors.forename: Yong
authors.initials: Y
authors.lastname: Tao
mesh_terms: D001711:Biotransformation; D004926:Escherichia coli / Q000201:enzymology*; D005822:Genetic Vectors; D014453:Glucuronosyltransferase / Q000235:genetics; D000076002:Hyaluronan Synthases; D006820:Hyaluronic Acid / Q000378:metabolism*; D016979:Pasteurella multocida / Q000201:enzymology; D013297:Streptococcus pyogenes / Q000201:enzymology*; D014533:Uridine Diphosphate Glucose Dehydrogenase / Q000378:metabolism*
chemical_list: D006820:Hyaluronic Acid; D014533:Uridine Diphosphate Glucose Dehydrogenase; D014453:Glucuronosyltransferase; D000076002:Hyaluronan Synthases