title: Production of taxadiene by engineering of mevalonate pathway in Escherichia coli and endophytic fungus Alternaria alternata TPF6.
abstract: Taxol (paclitaxel) is a diterpenoid compound with significant and extensive applications in the treatment of cancer. The production of Taxol and relevant intermediates by engineered microbes is an attractive alternative to the semichemical synthesis of Taxol. In this study, based on a previously developed platform, the authors first established taxadiene production in mutant E. coli T2 and T4 by engineering of the mevalonate (MVA) pathway. The authors then developed an Agrobacterium tumefaciens-mediated transformation (ATMT) method and verified the strength of heterologous promoters in Alternaria alternata TPF6. The authors next transformed the taxadiene-producing platform into A. alternata TPF6, and the MVA pathway was engineered, with introduction of the plant taxadiene-forming gene. Notably, by co-overexpression of isopentenyl diphosphate isomerase (Idi), a truncated version of 3-hydroxy-3-methylglutaryl-CoA reductase (tHMG1), and taxadiene synthase (TS), the authors could detect 61.9 ± 6.3 μg/L taxadiene in the engineered strain GB127. This is the first demonstration of taxadiene production in filamentous fungi, and the approach presented in this study provides a new method for microbial production of Taxol. The well-established ATMT method and the known promoter strengths facilitated further engineering of taxaenes in this fungus.
authors.affiliation: Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education and School of Pharmaceutical Sciences, Wuhan University, Wuhan, P. R. China.
authors.forename: Guangkai
authors.initials: G
authors.lastname: Bian
authors.affiliation: Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education and School of Pharmaceutical Sciences, Wuhan University, Wuhan, P. R. China.
authors.forename: Yujie
authors.initials: Y
authors.lastname: Yuan
authors.affiliation: Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education and School of Pharmaceutical Sciences, Wuhan University, Wuhan, P. R. China.
authors.forename: Hui
authors.initials: H
authors.lastname: Tao
authors.affiliation: Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education and School of Pharmaceutical Sciences, Wuhan University, Wuhan, P. R. China.
authors.forename: Xiaofei
authors.initials: X
authors.lastname: Shi
authors.affiliation: Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education and School of Pharmaceutical Sciences, Wuhan University, Wuhan, P. R. China.
authors.forename: Xiaofang
authors.initials: X
authors.lastname: Zhong
authors.affiliation: J1 Biotech Co., Ltd., Wuhan, P. R. China.
authors.forename: Yichao
authors.initials: Y
authors.lastname: Han
authors.affiliation: J1 Biotech Co., Ltd., Wuhan, P. R. China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Fu
authors.affiliation: China Center for Type Culture Collection (CCTCC), College of Life Sciences, Wuhan University, Wuhan, P. R. China.
authors.forename: Chengxiang
authors.initials: C
authors.lastname: Fang
authors.affiliation: Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education and School of Pharmaceutical Sciences, Wuhan University, Wuhan, P. R. China.|Hubei Engineering Laboratory for Synthetic Microbiology, Wuhan Institute of Biotechnology, Wuhan, P. R. China.
authors.forename: Zixin
authors.initials: Z
authors.lastname: Deng
authors.affiliation: Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education and School of Pharmaceutical Sciences, Wuhan University, Wuhan, P. R. China.|Hubei Engineering Laboratory for Synthetic Microbiology, Wuhan Institute of Biotechnology, Wuhan, P. R. China.
authors.forename: Tiangang
authors.initials: T
authors.lastname: Liu
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics / Q000378:metabolism; D000475:Alkenes / Q000378:metabolism* / Q000627:therapeutic use; D000528:Alternaria / Q000235:genetics / Q000378:metabolism; D019748:Carbon-Carbon Double Bond Isomerases / Q000096:biosynthesis; D004224:Diterpenes / Q000378:metabolism* / Q000627:therapeutic use; D060026:Endophytes / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D024243:HMGB1 Protein / Q000096:biosynthesis; D045782:Hemiterpenes; D006801:Humans; D007535:Isomerases / Q000096:biosynthesis; D060847:Metabolic Engineering; D008798:Mevalonic Acid / Q000378:metabolism*; D009369:Neoplasms / Q000188:drug therapy*; D014169:Transformation, Bacterial / Q000235:genetics
chemical_list: D000475:Alkenes; D004224:Diterpenes; D024243:HMGB1 Protein; C585491:HMGB1 protein, human; D045782:Hemiterpenes; C093125:taxa-4(5),11(12)diene; D007535:Isomerases; C095926:taxa-4(5),11(12)-diene synthase; D019748:Carbon-Carbon Double Bond Isomerases; C021453:isopentenyldiphosphate delta-isomerase; D008798:Mevalonic Acid
keywords: Agrobacterium tumefaciens; Alternaria alternata TPF6; Mevalonate pathway; Taxadiene