Abstract
Bacterial transporters mediate the exchanges between intracellular and extracellular environments. Modification of transport route could be applied to speed up the metabolic reactions and promote the production of aimed compounds. Herein, lysine 2-monooxygenase (DavB) and δ-aminovaleramidase (DavA) were co-expressed in Escherichia coli BL21(DE3) to produce nylon-5 monomer 5-aminovalerate from l-lysine. Then, PP2911 (4-aminobutyrate transporter in Pseudomonas putida) and LysP (the lysine specific permease in E. coli) were overexpressed to promote 5-aminovalerate production using whole cells of recombinant E. coli. The constructed E. coli strain overexpressing transport proteins exhibited good 5-aminovalerate production performance and might serve as a promising biocatalyst for 5-aminovalerate production from l-lysine. This strategy not only shows an efficient process for the production of nylon monomers but also might be used in production of other chemicals.
Highlights
(gene ID: 1044092) and davB of P. putida were inserted into the MCS1 of pETDuet-1 (Fig. S1A) to construct pETDuet-DavAB (Fig. S1B) which was transferred into E. coli BL21(DE3)
The inability of E. coli pD to consume l-lysine implied the suitability of E. coli as a host for efficient l-lysine conversion
L-lysine was consumed, no 5-aminovalerate was detected during the whole reaction process with E. coli pDB. These results suggested that both DavB and DavA should work efficiently in recombinant E. coli for the 5-aminovalerate production
Summary
Feasibility of 5-aminovalerate production by recombinant E. coli. Firstly, the genes davA (gene ID: 1044092) and davB (gene ID: 1044093) of P. putida were inserted into the MCS1 of pETDuet-1 (Fig. S1A) to construct pETDuet-DavAB (Fig. S1B) which was transferred into E. coli BL21(DE3). L-lysine was consumed, no 5-aminovalerate was detected during the whole reaction process with E. coli pDB These results suggested that both DavB and DavA should work efficiently in recombinant E. coli for the 5-aminovalerate production. In order to increase the export rate of 5-aminovalerate, the gene pp2911 (gene ID: 1042844) from P. putida KT2440 was inserted into pACYCDuet-1 (Fig. S1E) to construct pACYCDuet-PP2911 (Fig. S1F) It was transferred into E. coli pDAB to construct E. coli pDABP. The recombinant strain E. coli pDABLP successfully produced 16.9 g/L 5-aminovalerate from 22.5 g/L l-lysine in 24 h with a yield of 0.94 mol/mol (Table 1). Among all the constructed strains, the recombinant strain E. coli pDABLP overexpressing transport proteins LysP and PP2911 attained the best 5-aminovalerate production performance in titer, productivity and yield. The lower yield compared with batch biotransformation process might be caused by the instability of DavA, as mentioned in the previous research[18]
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