High-Effective Expression of the Gene for Aspartase (L-Aspartate Ammonium Lyase) in Escherichia coli Cells

Автор: A.D. Novikov, D.D. Derbikov, O.V. Shaposhnikova, T.A. Gubanova, S.V. Kameneva, and A.S. Yanenko

Страница: 19-24

High-Effective Expression of the Gene for Aspartase (L-Aspartate Ammonium Lyase) in Escherichia coli Cells

Russian Journal of Biotechnology, 2014, N 5, P. 19-24

UDC 577.214.622:579.66

Section:  “Producers, Biology, Selection, and Gene Engineering”

A.D. Novikov ^*,  D.D. Derbikov,  O.V. Shaposhnikova,  T.A. Gubanova,  S.V. Kameneva,  and  A.S. Yanenko

The State Research Institute for Genetics and Selection of Industrial Microorganisms,  117545, Moscow Russia

e-mail:  andrnov@genetika.ru

Cloning of the gene for aspartase (L-aspartase ammonium lyase) from a natural isolate of Escherichia coli VKPM B-7188 on a рLATE31 plasmid and its expression under the control of the T7 promoter in E. coli BLR(DE3) cells have been performed. It was shown that under the optimal conditions, the content of aspartase in the recombinant cells achieves 60% of the sum of soluble proteins. The maximum enzyme specific activity in the cells achieved 700 μmole/mg/min. The mutant variants bearing the C-end deleted aspartase genes with a higher level of the aspartase activity were obtained. The designed system of high-effective expression of the aspartase gene can be helpful in the selection and express-testing of catalytic characteristics of the in vitro generated mutant enzyme forms, and also in the design of more efficient biocatalysts for the L-aspartic acid synthesis.

Key words:  aspartase (L-aspartate-ammonium lyase),  cloning,  deleted gene variants,  Escherichia coli,  expression,  gene for aspartase.

24.03.2015, 1494 просмотра.