Gene Cloning，Expression and Enzyme Activity Assay of UDP-glucose Dehydrogenase in Escherichia coli

Abstract

Abstract: UDP-glucose dehydrogenase, as the key rate-limiting enzyme in the hyaluronic acid synthesis process, can catalyze UDP-glucose to UDP-glucuronic acid which is essential for cell to produce hyaluronic acid. The ugd gene amplified from genomic DNA of Escherichia coli BL21（DE3）and YK537 by PCR were cloned into pBLMVL2 vector containing P _{L promoter. The plasmid pBLBugd and pBLYugd were constructed and transformed respectively into Escherichia coli BL21（DE3）and YK537 to get four recombinant strains. UDP-glucose dehydrogenase activity of different recombinant strains was measured by thermal induction. The differences of expression level and enzyme activity of UDP-glucose dehydrogenase were detected under different temperature induced conditions. These result demonstrate that adopting double stage of heating induction and adding some moderate amount of yeast extract to the cultivating medium can improve the expression level and enzyme activity of UDP - glucose dehydrogenase.}

Key words: Hyaluronic acid, UDP-glucose dehydrogenase, P_L promoter, Gene sequencing, Enzyme activity

Chen Yihan,Qian Yue,Hou Yongtai,Zhou Qingwei,Gan Renbao,Guan Shimin,Rong Shaofeng. Gene Cloning，Expression and Enzyme Activity Assay of UDP-glucose Dehydrogenase in Escherichia coli[J]. Biotechnology Bulletin, 2013, 0(7): 136-143.

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