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Unexpected NADPH Hydratase Activity in the Nitrile Reductase QueF from Escherichia coli

The nitrile reductase QueF catalyzes NADPH‐dependent reduction of the nitrile group of preQ(0) (7‐cyano‐7‐deazaguanine) into the primary amine of preQ(1) (7‐aminomethyl‐7‐deazaguanine), a biologically unique reaction important in bacterial nucleoside biosynthesis. Here we have discovered that the Qu...

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Autores principales: Jung, Jihye, Braun, Jan, Czabany, Tibor, Nidetzky, Bernd
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7317782/
https://www.ncbi.nlm.nih.gov/pubmed/31850614
http://dx.doi.org/10.1002/cbic.201900679
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author Jung, Jihye
Braun, Jan
Czabany, Tibor
Nidetzky, Bernd
author_facet Jung, Jihye
Braun, Jan
Czabany, Tibor
Nidetzky, Bernd
author_sort Jung, Jihye
collection PubMed
description The nitrile reductase QueF catalyzes NADPH‐dependent reduction of the nitrile group of preQ(0) (7‐cyano‐7‐deazaguanine) into the primary amine of preQ(1) (7‐aminomethyl‐7‐deazaguanine), a biologically unique reaction important in bacterial nucleoside biosynthesis. Here we have discovered that the QueF from Escherichia coli—its D197A and E89L variants in particular (apparent k (cat)≈10(−2) min(−1))—also catalyze the slow hydration of the C5=C6 double bond of the dihydronicotinamide moiety of NADPH. The enzymatically C6‐hydrated NADPH is a 3.5:1 mixture of R and S forms and rearranges spontaneously through anomeric epimerization (β→α) and cyclization at the tetrahydronicotinamide C6 and the ribosyl O2. NADH and 1‐methyl‐ or 1‐benzyl‐1,4‐dihydronicotinamide are not substrates of the enzymatic hydration. Mutagenesis results support a QueF hydratase mechanism, in which Cys190—the essential catalytic nucleophile for nitrile reduction—acts as the general acid for protonation at the dihydronicotinamide C5 of NADPH. Thus, the NADPH hydration in the presence of QueF bears mechanistic resemblance to the C=C double bond hydration in natural hydratases.
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spelling pubmed-73177822020-06-29 Unexpected NADPH Hydratase Activity in the Nitrile Reductase QueF from Escherichia coli Jung, Jihye Braun, Jan Czabany, Tibor Nidetzky, Bernd Chembiochem Full Papers The nitrile reductase QueF catalyzes NADPH‐dependent reduction of the nitrile group of preQ(0) (7‐cyano‐7‐deazaguanine) into the primary amine of preQ(1) (7‐aminomethyl‐7‐deazaguanine), a biologically unique reaction important in bacterial nucleoside biosynthesis. Here we have discovered that the QueF from Escherichia coli—its D197A and E89L variants in particular (apparent k (cat)≈10(−2) min(−1))—also catalyze the slow hydration of the C5=C6 double bond of the dihydronicotinamide moiety of NADPH. The enzymatically C6‐hydrated NADPH is a 3.5:1 mixture of R and S forms and rearranges spontaneously through anomeric epimerization (β→α) and cyclization at the tetrahydronicotinamide C6 and the ribosyl O2. NADH and 1‐methyl‐ or 1‐benzyl‐1,4‐dihydronicotinamide are not substrates of the enzymatic hydration. Mutagenesis results support a QueF hydratase mechanism, in which Cys190—the essential catalytic nucleophile for nitrile reduction—acts as the general acid for protonation at the dihydronicotinamide C5 of NADPH. Thus, the NADPH hydration in the presence of QueF bears mechanistic resemblance to the C=C double bond hydration in natural hydratases. John Wiley and Sons Inc. 2020-02-20 2020-05-15 /pmc/articles/PMC7317782/ /pubmed/31850614 http://dx.doi.org/10.1002/cbic.201900679 Text en © 2019 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA. This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.
spellingShingle Full Papers
Jung, Jihye
Braun, Jan
Czabany, Tibor
Nidetzky, Bernd
Unexpected NADPH Hydratase Activity in the Nitrile Reductase QueF from Escherichia coli
title Unexpected NADPH Hydratase Activity in the Nitrile Reductase QueF from Escherichia coli
title_full Unexpected NADPH Hydratase Activity in the Nitrile Reductase QueF from Escherichia coli
title_fullStr Unexpected NADPH Hydratase Activity in the Nitrile Reductase QueF from Escherichia coli
title_full_unstemmed Unexpected NADPH Hydratase Activity in the Nitrile Reductase QueF from Escherichia coli
title_short Unexpected NADPH Hydratase Activity in the Nitrile Reductase QueF from Escherichia coli
title_sort unexpected nadph hydratase activity in the nitrile reductase quef from escherichia coli
topic Full Papers
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7317782/
https://www.ncbi.nlm.nih.gov/pubmed/31850614
http://dx.doi.org/10.1002/cbic.201900679
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