Angelastro, Antonio ![]() ![]() ![]() ![]() |
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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 |
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Date Type: | Publication |
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: | 05 May 2023 07:30 |
URI: | https://orca.cardiff.ac.uk/id/eprint/97525 |
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