Enhancing isoprenoid production through systematically assembling and modulating efflux pumps in Escherichia coli
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Enhancement of the cellular exportation of heterologous compounds is an important aspect to improve the product yield in microbial cell factory. Efflux pumps can expel various intra- or extra-cellular substances out of microbial hosts and increase the cellular tolerance. Thus in this study, by using the hydrophobic sesquiterpene (amorphadiene) and diterpene (kaurene) as two model compounds, we attempted to improve isoprenoid production through systematically engineering the efflux pumps in Escherichia coli BL21(DE3). The pleiotropic resistant pumps, AcrAB-TolC, MdtEF-TolC from E. coli and heterologous MexAB-OprM pump from Pseudomonas aeruginosa, were overexpressed, assembled, and finely modulated. We found that overexpression of AcrB and TolC components can effectively enhance the specific yield of amorphadiene and kaurene, e.g., 31 and 37 % improvement for amorphadiene compared with control, respectively. The heterologous MexB component can enhance kaurene production with 70 % improvement which is more effective than TolC and AcrB. The results suggest that the three components of tripartite efflux pumps play varied effect to enhance isoprenoid production. Considering the highly organized structure of efflux pumps and importance of components interaction, various component combinations were constructed and the copy number of key components AcrB and TolC was finely modulated as well. The results exhibit that the combination TolC and TolC and AcrB improved the specific yield of amorphadiene with 118 %, and AcrA and TolC and AcrB improved that of kaurene with 104 %. This study indicates that assembling and finely modulating efflux pumps is an effective strategy to improve the production of heterologous compounds in E. coli.
KeywordsEfflux pumps Heterologous production Escherichia coli Isoprenoids Amorphadiene Kaurene
This work was supported by the National Basic Research Program of China (“973” Program, grant no. 2012CB721104), the National High Technology Research and Development Program (“863” Program, grant no. 2012AA02A701), the National Natural Science Foundation of China (grants no. 31170101 and 31100073), “Technology Innovation Action Plan” Key Project of Shanghai Science and Technology Commission (grant no. 10dz1910100), and the major Projects of Knowledge Innovation Program of Chinese Academy of Sciences (grant no. KSCX2-EW-J-12).
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