A ‘Split-Gene’ Transketolase From the Hyper-Thermophilic Bacterium Carboxydothermus hydrogenoformans: Structure and Biochemical Characterization

James, Paul, Isupov, Michail N., De Rose, Simone Antonio, Sayer, Christopher, Cole, Isobel S. and Littlechild, Jennifer A. (2020) A ‘Split-Gene’ Transketolase From the Hyper-Thermophilic Bacterium Carboxydothermus hydrogenoformans: Structure and Biochemical Characterization. Frontiers in Microbiology, 11. p. 592353. ISSN 1664-302X

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Official URL: https://doi.org/10.3389/fmicb.2020.592353

Abstract

A novel transketolase has been reconstituted from two separate polypeptide chains encoded by a 'split-gene' identified in the genome of the hyperthermophilic bacterium, Carboxydothermus hydrogenoformans. The reconstituted active α2β2 tetrameric enzyme has been biochemically characterized and its activity has been determined using a range of aldehydes including glycolaldehyde, phenylacetaldehyde and cyclohexanecarboxaldehyde as the ketol acceptor and hydroxypyruvate as the donor. This reaction proceeds to near 100% completion due to the release of the product carbon dioxide and can be used for the synthesis of a range of sugars of interest to the pharmaceutical industry. This novel reconstituted transketolase is thermally stable with no loss of activity after incubation for 1 h at 70°C and is stable after 1 h incubation with 50% of the organic solvents methanol, ethanol, isopropanol, DMSO, acetonitrile and acetone. The X-ray structure of the holo reconstituted α2β2 tetrameric transketolase has been determined to 1.4 Å resolution. In addition, the structure of an inactive tetrameric β4 protein has been determined to 1.9 Å resolution. The structure of the active reconstituted α2β2 enzyme has been compared to the structures of related enzymes; the E1 component of the pyruvate dehydrogenase complex and D-xylulose-5-phosphate synthase, in an attempt to rationalize differences in structure and substrate specificity between these enzymes. This is the first example of a reconstituted 'split-gene' transketolase to be biochemically and structurally characterized allowing its potential for industrial biocatalysis to be evaluated.

Item Type: Article
Uncontrolled Keywords: hyperthermophilic, ‘split-gene’, transketolase, thermal stability, industrial applications
Subjects: C500 Microbiology
Department: Faculties > Health and Life Sciences > Applied Sciences
Depositing User: John Coen
Date Deposited: 19 Nov 2020 14:26
Last Modified: 19 Nov 2020 14:30
URI: http://nrl.northumbria.ac.uk/id/eprint/44797

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