title: Enhanced pinocembrin production in Escherichia coli by regulating cinnamic acid metabolism.
abstract: Microbial biosynthesis of pinocembrin is of great interest in the area of drug research and human healthcare. Here we found that the accumulation of the pathway intermediate cinnamic acid adversely affected pinocembrin production. Hence, a stepwise metabolic engineering strategy was carried out aimed at eliminating this pathway bottleneck and increasing pinocembrin production. The screening of gene source and the optimization of gene expression was first employed to regulate the synthetic pathway of cinnamic acid, which showed a 3.53-fold increase in pinocembrin production (7.76 mg/L) occurred with the alleviation of cinnamic acid accumulation in the engineered E. coli. Then, the downstream pathway that consuming cinnamic acid was optimized by the site-directed mutagenesis of chalcone synthase and cofactor engineering. S165M mutant of chalcone synthase could efficiently improve the pinocembrin production, and allowed the product titer of pinocembrin increased to 40.05 mg/L coupled with the malonyl-CoA engineering. With a two-phase pH fermentation strategy, the cultivation of the optimized strain resulted in a final pinocembrin titer of 67.81 mg/L. The results and engineering strategies demonstrated here would hold promise for the titer improvement of other flavonoids.
authors: State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing 211816, P.R. China.|College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, P.R. China.
authors: Weijia
authors: W
authors: Cao
authors: State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing 211816, P.R. China.|College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, P.R. China.|College of Bioengineering and Biotechnology, Tianshui Normal University, Tianshui 741001, P.R. China.
authors: Weichao
authors: W
authors: Ma
authors: State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing 211816, P.R. China.|College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, P.R. China.
authors: Xin
authors: X
authors: Wang
authors: State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing 211816, P.R. China.|College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, P.R. China.
authors: Bowen
authors: B
authors: Zhang
authors: State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing 211816, P.R. China.|College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, P.R. China.
authors: Xun
authors: X
authors: Cao
authors: State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing 211816, P.R. China.|College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, P.R. China.
authors: Kequan
authors: K
authors: Chen
authors: State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing 211816, P.R. China.|College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, P.R. China.
authors: Yan
authors: Y
authors: Li
authors: State Key Laboratory of Materials-Oriented Chemical Engineering, Nanjing 211816, P.R. China.|College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, P.R. China.
authors: Pingkai
authors: P
authors: Ouyang
mesh_terms: D000217:Acyltransferases / Q000378:metabolism; D061249:Batch Cell Culture Techniques; D018533:Biomass; D002934:Cinnamates / Q000378:metabolism*; D004926:Escherichia coli / Q000187:drug effects / Q000378:metabolism*; D005285:Fermentation / Q000187:drug effects; D044950:Flavanones / Q000096:biosynthesis* / Q000737:chemistry; D018628:Gene Dosage; D015870:Gene Expression / Q000187:drug effects; D005947:Glucose / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008316:Malonyl Coenzyme A / Q000378:metabolism; D060847:Metabolic Engineering; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D013997:Time Factors
chemical_list: D002934:Cinnamates; D044950:Flavanones; C029010:cinnamic acid; D010649:Phenylalanine; D008316:Malonyl Coenzyme A; C016063:pinocembrin; D000217:Acyltransferases; C020141:flavanone synthetase; D005947:Glucose