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A versatile disulfide-driven recycling system for NADP+ with high cofactor turnover number

Angelastro, Antonio, Dawson, William M., Luk, Louis and Allemann, Rudolf 2016. A versatile disulfide-driven recycling system for NADP+ with high cofactor turnover number. ACS Catalysis 7 , pp. 1025-1029. 10.1021/acscatal.6b03061

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Abstract

NADP+-dependent enzymes are important in many biocatalytic processes to generate high-value chemicals for the pharmaceutical and food industry; hence, a costeffective, efficient, and environmentally friendly recycling system for the relatively expensive and only marginally stable enzyme cofactor NADP+ offers significant benefits. NADP+ regeneration schemes have previously been described, but their application is severely limited by the low total turnover numbers (TTN) for the cofactor. Here, we report a glutathione-based recycling system that combines glutaredoxin from E. coli (EcGRX) and the glutathione reductase from S. cerevisiae (ScGR) for NADP+ regeneration. This system employs inexpensive latent organic disulfides such as oxidized cysteine or 2-hydroxyethyl disulfide (HED) as oxidizing agents and allows NADP+ recycling under both aerobic and anaerobic conditions with a TTN in excess of 5 × 105, indicating that each regeneration cycle is 99.9998% selective toward forming the cofactor. Accordingly, for each 1 mol of product generated, less than $0.05 of cofactor is needed. Finally, the EcGRX/ScGR pair is compatible with eight enzymes in the guanosine monophosphate (GMP) biosynthetic pathway, giving the corresponding isotopically labeled nucleotide in high yield. The glutathione-based NADP+ recycling system has potential for biocatalytic applications in academic and industrial settings.

Item Type: Article
Date Type: Published Online
Status: Published
Schools: Chemistry
Subjects: Q Science > QD Chemistry
Uncontrolled Keywords: biocatalysis, biosynthesis, cofactor/coenzyme recycling, enzyme oxidation, biotechnology
Publisher: American Chemical Society
ISSN: 2155-5435
Funders: Biotechnology and Biological Sciences Research Council
Date of First Compliant Deposit: 18 January 2017
Date of Acceptance: 19 December 2016
Last Modified: 06 Sep 2017 09:04
URI: http://orca.cf.ac.uk/id/eprint/97525

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