Engineering a nicotinamide mononucleotide redox cofactor system for biocatalysis
Biological production of chemicals often requires the use of cellular cofactors, such as nicotinamide adenine dinucleotide phosphate (NADP + ). These cofactors are expensive to use in vitro and difficult to control in vivo. We demonstrate the development of a noncanonical redox cofactor system based...
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Published in | Nature chemical biology Vol. 16; no. 1; pp. 87 - 94 |
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Main Authors | , , , , , , , , , |
Format | Journal Article |
Language | English |
Published |
New York
Nature Publishing Group US
01.01.2020
Nature Publishing Group |
Subjects | |
Online Access | Get full text |
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Summary: | Biological production of chemicals often requires the use of cellular cofactors, such as nicotinamide adenine dinucleotide phosphate (NADP
+
). These cofactors are expensive to use in vitro and difficult to control in vivo. We demonstrate the development of a noncanonical redox cofactor system based on nicotinamide mononucleotide (NMN
+
). The key enzyme in the system is a computationally designed glucose dehydrogenase with a 10
7
-fold cofactor specificity switch toward NMN
+
over NADP
+
based on apparent enzymatic activity. We demonstrate that this system can be used to support diverse redox chemistries in vitro with high total turnover number (~39,000), to channel reducing power in
Escherichia coli
whole cells specifically from glucose to a pharmaceutical intermediate, levodione, and to sustain the high metabolic flux required for the central carbon metabolism to support growth. Overall, this work demonstrates efficient use of a noncanonical cofactor in biocatalysis and metabolic pathway design.
Redesign of a glucose dehydrogenase to use nicotinamide mononucleotide (NMN
+
) instead of NAD(P)
+
enables the development of a noncanonical cofactor system that can be used to support redox chemistries both in vitro and in
Escherichia coli
. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 H.L. and J.B.S. conceived the research. L.Z., W.B.B., and E.K. performed mutant enzyme kinetics characterization. W.B.B., L.Z., S.M., E.K., and B.F. performed the in vitro biotransformation. W.B.B., S.M., B.F., and A.S.M. performed the whole-cell biotransformation. W.B.B. performed the intracelluar NMN+ and NAD+ level analysis. S.M. performed the NMN+-dependent growth experiments. W.S.M. and Y.C. performed the computational modeling. All authors analyzed the data and wrote the manuscript. Author contributions |
ISSN: | 1552-4450 1552-4469 |
DOI: | 10.1038/s41589-019-0402-7 |