期刊
JOURNAL OF BIOTECHNOLOGY
卷 289, 期 -, 页码 135-143出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jbiotec.2018.11.010
关键词
NADH oxidase; Co-factor regeneration; Lactobacillus reuteri; 3-Hydroxypropionic acid; 3-Hydroxypropionaldehyde; Fed-batch biotransformation
资金
- Vinnova (The Swedish Innovation Agency) [2008-00849]
- National Council for Research of Brazil (Conselho Nacional de Pesquisa, CNPQ)
- Forte [2008-00849] Funding Source: Forte
- Formas [2008-00849] Funding Source: Formas
- Vinnova [2008-00849] Funding Source: Vinnova
Lactobacillus reuteri metabolizes glycerol through propanediol-utilization (Pdu) pathway to 1,3-propanediol (1,3PD) via 3-hydroxypropionaldehyde (3-HPA) as intermediate. In the resting cells, the oxidized co-factor obtained in the reaction is regenerated by simultaneous oxidation of 3-HPA to 3-hydroxypropionic acid (3-HP) using propionaldehyde dehydrogenase (PduP), phosphotransacylase (PduL) and propionate kinase (PduW). We have earlier shown that the use of resting cells of recombinant Escherichia coli expressing the oxidative pathway gives the highest theoretical yield of 1 mol 3-HP per mol 3-HPA but is limited by cofactor depletion. In the present study, the gene encoding the enzyme NAD(P) H oxidase (LreuNox) that utilizes molecular oxygen as substrate, was isolated from L. reuteri and heterologously overexpressed in E. coli. LreuNox has a pH optimum of 6 and exhibits V-max of 101.1 +/- 2.2 U/ mg with NADH, which is 30% higher than that for NADPH. Co-expression of LreuNox with PduP, PduL and PduW in E. coli enhances the biocatalytic lifetime as well as productivity at least two-fold compared to that achieved without co-factor regeneration.
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