Enhanced production of coenzyme Q10 by self-regulating the engineered MEP pathway in Rhodobacter sphaeroides

• Published in: Biotechnology and bioengineering Vol. 111; no. 4; pp. 761 - 769
• Main Authors: Lu, Wenqiang, Ye, Lidan, Xu, Haoming, Xie, Wenping, Gu, Jiali, Yu, Hongwei
• Format: Journal Article
• Language: English
• Published: United States Blackwell Publishing Ltd 01.04.2014
• Subjects: coenzyme Q10; constitutive expression; Erythritol - analogs & derivatives; Erythritol - metabolism; LacIq; metabolic balance; Metabolic Engineering - methods; Metabolic Networks and Pathways - genetics; Metabolic Networks and Pathways - physiology; Rhodobacter sphaeroides - genetics; Rhodobacter sphaeroides - metabolism; Rhodobacter sphaeroides - physiology; ribosomal bind site; self-regulation; Ubiquinone - analogs & derivatives; Ubiquinone - analysis; Ubiquinone - metabolism; ribosomal bind site; constitutive expression; self-regulation; coenzyme Q10; LacIq; metabolic balance
• ISSN: 0006-3592; 1097-0290; 1097-0290
• DOI: 10.1002/bit.25130

ABSTRACT Fine‐tuning the expression level of an engineered pathway is crucial for the metabolic engineering of a host toward a desired phenotype. However, most engineered hosts suffer from nonfunctional protein expression, metabolic imbalance, cellular burden or toxicity from intermediates when an engineered pathway is first introduced, which can decrease production of the desired product. To circumvent these obstacles, we developed a self‐regulation system utilizing the trc/tac promoter, LacIq protein and ribosomal binding sites (RBS). With the purpose of improving coenzyme Q10 (CoQ10) production by increasing the decaprenyl diphosphate supplement, enzymes DXS, DXR, IDI, and IspD were constitutively overexpressed under the control of the trc promoter in Rhodobacter sphaeroides. Then, a self‐regulation system combining a set of RBSs for adjusting the expression of the LacIq protein was applied to tune the expression of the four genes, resulting in improved CoQ10 production. Finally, another copy of the tac promoter with the UbiG gene (involved in the ubiquinone pathway of CoQ10 biosynthesis) was introduced into the engineered pathway. By optimizing the expression level of both the upstream and downstream pathway, CoQ10 production in the mutants was improved up to 93.34 mg/L (7.16 mg/g DCW), about twofold of the wild‐type (48.25 mg/L, 3.24 mg/g DCW). Biotechnol. Bioeng. 2014;111: 761–769. © 2013 Wiley Periodicals, Inc. A self‐regulating system has been developed to control the expression level of the engineered pathway via the co‐expressed LacIq protein. This system could flexibly regulate the engineered pathway to balance with the host metabolic pathways. With this system, the MEP pathway in Rhodobacter sphaeroides was optimized and the production of coenzyme Q10 was improved two‐folds.