Gene cloning and expression of a glucoside 3-dehydrogenase from Sphingobacterium faecium ZJF-D6, and used it to produce N-p-nitrophenyl-3-ketovalidamine

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• 作者：Jian-Fen Zhang ; Wei-Qing Chen ; Hong Chen
• 关键词：Glucoside 3 ; dehydrogenase ; 3 ; Ketovalidoxylamine A C–N lyase ; N ; p ; nitrophenyl ; 3 ; ketovalidamine ; Valienamine ; Gene cloning
• 刊名：World Journal of Microbiology and Biotechnology
• 出版年：2017
• 出版时间：February 2017
• 年：2017
• 卷：33
• 期：2
• 全文大小：
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Applied Microbiology; Biotechnology; Biochemistry, general; Environmental Engineering/Biotechnology; Microbiology;
• 出版者：Springer Netherlands
• ISSN：1573-0972
• 卷排序：33

文摘

In this study, we report the cloning and expression of a functional glucoside 3-dehydrogenase (G3DH) gene from Sphingobacterium faecium ZJF-D6. This gene is 1686 bp in length and encodes a peptide of 562 amino acids. The G3DH gene was successfully expressed in E. coli, and the recombinant enzyme could oxidize glucosides, galactosides and analogues at C-3 position. The sequence and multiple alignment analysis showed that the enzyme has highest identity with G3DHs from Paraglaciecola polaris LMG 21857, Aliiglaciecola lipolytica E3 and Halomonas sp. alpha-15. The recombinant G3DH was purified on Ni–NTA column and exhibited the highest activity at pH 7.6 and 30 °C. It was sensitive to acid and alkali, and showed well thermostability. The SfG3DH could oxidize a wild range of sugars. When recombinant E. coli BL21 cells were used as catalyst, a high rate of conversion to N-p-nitrophenyl-3-ketovalidamine was achieved, and no p-nitroaniline was detected. This process offers a promising approach to fulfill substrate of 3-ketovalidoxylamine A C–N lyase production.