Study on Aerobic Biosynthesis of 4-Hydroxybutyric Acid by Escherichia coli Cells with Heterological Expression of 2-Ketoglutarate Decarboxylase Gene

Автор: A.Yu. Gulevich, M.S. Skonechnyi, A.V. Sukhozhenko, A.Yu. Skorokhodova, and V.G. Debabov

Страница: 46-54

Study on Aerobic Biosynthesis of 4-Hydroxybutyric Acid by Escherichia coli Cells with Heterological Expression of 2-Ketoglutarate Decarboxylase Gene

Biotekhnologiya, 2015, N 2, P. 46-54

UDC 577.121

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

A.Yu. Gulevich ^{1, *},  M.S. Skonechnyi ²,  A.V. Sukhozhenko ¹,  A.Yu. Skorokhodova ¹,  and  V.G. Debabov ¹

¹  The State Research Institute for Genetics and Selection of Industrial Microorganisms (GosNIIgenetika),  117545, Moscow Russia

²  The Russian University of People’s Friendship,  117198, Moscow Russia

e-mail:  gulevich@genetika.ru

The Mycobacterium tuberculosis Rv1248c(kgd) gene has been expressed in the recombinant Escherichia coli strain with the inactivated pathways of mixed-acid fermentation and aerobic generation of acetyl-CoA, and also with modified systems of glucose transport and phosphorylation, and gene ydfC regulation (the latter encoding the NADPH- dependant dehydrogenase of hydrocarboxylic acids). It was established that on the background of the intense formation of 2-ketoglutarate during the glucose aerobic utilization, the direct conversion of 2-ketoglutarate to succinate semialdehyde under the action of an exogenous enzymatic activity is not the only pathway for the synthesis of 4-hydroxybutyric acid by the recombinant E. coli strain. This process can also proceed via the involvement of the appropriate intermediate of the TCA cycle in a cascade of native biochemical reactions. The induced 2-ketoglutarate-decarboxylase gene expression in the recombinant strain provided the efficient conversion of 2-ketoglutarate to succinate semialdehyde derivatives which ensured the yield of the synthesized 4-hydroxybutyric acid up to 0.3 mM and seemed to be limited by the activity of the enzyme responsible for the terminal stage of the precursor reduction.

Key words:  Escherichia coli,  4-hydroxybutyric acid,  2-ketoglutarate decarboxylase.

25.06.2015, 1422 просмотра.