title: Directed accumulation of less toxic pimaricin derivatives by improving the efficiency of a polyketide synthase dehydratase domain.
abstract: Pimaricin is an important polyene antifungal antibiotic that binds ergosterol and extracts it from fungal membranes. In previous work, two pimaricin derivatives (1 and 2) with improved pharmacological activities and another derivative (3) that showed no antifungal activity were produced by the mutant strain of Streptomyces chattanoogensis, in which the P450 monooxygenase gene scnG has been inactivated. Furthermore, inactivation of the DH12 dehydratase domain of the pimaricin polyketide synthases (PKSs) resulted in specific accumulation of the undesired metabolite 3, suggesting that improvement of the corresponding dehydratase activity may reduce or eliminate the accumulation of 3. Accordingly, the DH12-KR12 didomain within the pimaricin PKS was swapped with the fully active DH11-KR11 didomain. As predicted, the mutant was not able to produce 3 but accumulated 1 and 2 in high yields. Moreover, the effect of the flanking linker regions on domain swapping was evaluated. It was found that retention of the DH12-KR12 linker regions was more critical for the processivity of hybrid PKSs. Subsequently, high-yield production of 1 or 2 was obtained by overexpressing the scnD gene and its partner scnF and by disrupting the scnD gene, respectively. To our knowledge, this is the first report on the elimination of a polyketide undesired metabolite along with overproduction of desired product by improving the catalytic efficiency of a DH domain using a domain swapping technology.
authors.affiliation: State Key Laboratory of Microbial Metabolism, and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China.|Joint International Research Laboratory of Metabolic and Developmental Sciences, Shanghai Jiao Tong University, Shanghai, 200240, China.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Qi
authors.affiliation: State Key Laboratory of Microbial Metabolism, and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China.|Joint International Research Laboratory of Metabolic and Developmental Sciences, Shanghai Jiao Tong University, Shanghai, 200240, China.
authors.forename: Yucong
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Microbial Metabolism, and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China.|Joint International Research Laboratory of Metabolic and Developmental Sciences, Shanghai Jiao Tong University, Shanghai, 200240, China.
authors.forename: Qianjin
authors.initials: Q
authors.lastname: Kang
authors.affiliation: State Key Laboratory of Microbial Metabolism, and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China.|Joint International Research Laboratory of Metabolic and Developmental Sciences, Shanghai Jiao Tong University, Shanghai, 200240, China.
authors.forename: Chunyan
authors.initials: C
authors.lastname: Jiang
authors.affiliation: State Key Laboratory of Microbial Metabolism, and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China.|Joint International Research Laboratory of Metabolic and Developmental Sciences, Shanghai Jiao Tong University, Shanghai, 200240, China.
authors.forename: Jianting
authors.initials: J
authors.lastname: Zheng
authors.affiliation: State Key Laboratory of Microbial Metabolism, and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China. bailq@sjtu.edu.cn.|Joint International Research Laboratory of Metabolic and Developmental Sciences, Shanghai Jiao Tong University, Shanghai, 200240, China. bailq@sjtu.edu.cn.
authors.forename: Linquan
authors.initials: L
authors.lastname: Bai
mesh_terms: D000935:Antifungal Agents / Q000737:chemistry / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D004875:Ergosterol / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D009154:Mutation; D010866:Natamycin / Q000096:biosynthesis* / Q000737:chemistry; D048630:Polyketide Synthases / Q000235:genetics* / Q000378:metabolism; D000072417:Protein Domains; D015202:Protein Engineering; D013302:Streptomyces / Q000235:genetics* / Q000378:metabolism; D013329:Structure-Activity Relationship
chemical_list: D000935:Antifungal Agents; D001426:Bacterial Proteins; D048630:Polyketide Synthases; D010866:Natamycin; D004875:Ergosterol
keywords: Component elimination; Domain swapping; Pimaricin; Polyene macrolide; Polyketide