Journal
SYNTHETIC AND SYSTEMS BIOTECHNOLOGY
Volume 7, Issue 1, Pages 513-521Publisher
KEAI PUBLISHING LTD
DOI: 10.1016/j.synbio.2021.12.008
Keywords
L-phenylglycine; Whole-cell biocatalyst; Self-sufficient; Protein scaffold; Hydroxymandelate synthase
Categories
Funding
- National Natural Science Foundation of China [31900064]
- Natural Science Foundation of Heilongjiang Province of China [LH2019C012]
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An L-phg synthesis pathway was developed in Escherichia coli, resulting in increased L-phg production by introducing new enzymes, a cofactor self-sufficient system, and a protein scaffold.
L-phenylglycine (L-phg) is a valuable non-proteinogenic amino acid used as a precursor to beta-lactam antibiotics, antitumor agent taxol and many other pharmaceuticals. L-phg synthesis through microbial bioconversion allows for high enantioselectivity and sustainable production, which will be of great commercial and environmental value compared with organic synthesis methods. In this work, an L-phg synthesis pathway was built in Escherichia coli resulting in 0.23 mM L-phg production from 10 mM L-phenylalanine. Then, new hydroxymandelate synthases and hydroxymandelate oxidases were applied in the L-phg synthesis leading to a 5-fold increase in L-phg production. To address 2-oxoglutarate, NH4+, and NADH shortage, a cofactor self-sufficient system was introduced, which converted by-product L-glutamate and NAD(+) to these three cofactors simultaneously. In this way, L-phg increased 2.5-fold to 2.82 mM. Additionally, in order to reduce the loss of these three cofactors, a protein scaffold between synthesis pathway and cofactor regeneration modular was built, which further improved the L-phg production to 3.72 mM with a yield of 0.34 g/g L-phe. This work illustrated a strategy applying for whole-cell biocatalyst converting amino acid to its value-added chiral amine in a cofactor selfsufficient manner.
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