title: Complete elucidation of the late steps of bafilomycin biosynthesis inStreptomyces lohii.
abstract: Bafilomycins are an important subgroup of polyketides with diverse biological activities and possible applications as specific inhibitors of vacuolar H+-ATPase. However, the general toxicity and structural complexity of bafilomycins present formidable challenges to drug design via chemical modification, prompting interests in improving bafilomycin activities via biosynthetic approaches. Two bafilomycin biosynthetic gene clusters have been identified, but their post-polyketide synthase (PKS) tailoring steps for structural diversification and bioactivity improvement remain largely unknown. In this study, the post-PKS tailoring pathway from bafilomycin A1(1)→C1(2)→B1(3) in the marine microorganismStreptomyces lohiiwas elucidated for the first time byin vivogene inactivation andin vitrobiochemical characterization. We found that fumarate is first adenylated by a novel fumarate adenylyltransferase Orf3. Then, the fumaryl transferase Orf2 is responsible for transferring the fumarate moiety from fumaryl-AMP to the 21-hydroxyl group of 1 to generate 2. Last, the ATP-dependent amide synthetase BafY catalyzes the condensation of 2 and 2-amino-3-hydroxycyclopent-2-enone (C5N) produced by the 5-aminolevulinic acid synthase BafZ and the acyl-CoA ligase BafX, giving rise to the final product 3. The elucidation of fumarate incorporation mechanism represents the first paradigm for biosynthesis of natural products containing the fumarate moiety. Moreover, the bafilomycin post-PKS tailoring pathway features an interesting cross-talk between primary and secondary metabolisms for natural product biosynthesis. Taken together, this work provides significant insights into bafilomycin biosynthesis to inform future pharmacological development of these compounds.
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101.|the University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Zhong
authors.initials: Z
authors.lastname: Li
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101.|the University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Du
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101.
authors.forename: Xingwang
authors.initials: X
authors.lastname: Zhang
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101.|the University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Jiang
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101.
authors.forename: Kun
authors.initials: K
authors.lastname: Liu
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101.
authors.forename: Ping
authors.initials: P
authors.lastname: Men
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101.
authors.forename: Huifang
authors.initials: H
authors.lastname: Xu
authors.affiliation: the Departments of Medicinal Chemistry, Chemistry, and Microbiology and Immunology, Life Sciences Institute, University of Michigan, Ann Arbor, Michigan 48109, and.
authors.forename: Jeffrey L
authors.initials: JL
authors.lastname: Fortman
authors.affiliation: the Departments of Medicinal Chemistry, Chemistry, and Microbiology and Immunology, Life Sciences Institute, University of Michigan, Ann Arbor, Michigan 48109, and.
authors.forename: David H
authors.initials: DH
authors.lastname: Sherman
authors.affiliation: the State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, Guangdong 510060, China.
authors.forename: Bing
authors.initials: B
authors.lastname: Yu
authors.affiliation: the State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, Guangdong 510060, China.
authors.forename: Song
authors.initials: S
authors.lastname: Gao
authors.affiliation: From the Shandong Provincial Key Laboratory of Synthetic Biology, and CAS Key Laboratory of Biofuels at Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, Shandong 266101, lishengying@qibebt.ac.cn.
authors.forename: Shengying
authors.initials: S
authors.lastname: Li
mesh_terms: D000624:5-Aminolevulinate Synthetase / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000737:chemistry; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D015195:Drug Design; D005650:Fumarates / Q000737:chemistry; D005798:Genes, Bacterial; D005822:Genetic Vectors; D007700:Kinetics; D018942:Macrolides / Q000737:chemistry*; D005810:Multigene Family; D016366:Open Reading Frames; D048630:Polyketide Synthases / Q000378:metabolism; D061065:Polyketides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D013302:Streptomyces / Q000378:metabolism*; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism
chemical_list: D005650:Fumarates; D018942:Macrolides; D061065:Polyketides; C057620:bafilomycin A; D048630:Polyketide Synthases; C052501:bafilomycin B1; C052502:bafilomycin C1; D000255:Adenosine Triphosphate; D000624:5-Aminolevulinate Synthetase; D025262:Vacuolar Proton-Translocating ATPases
keywords: antibiotics; microbiology; natural product biosynthesis; polyketide; secondary metabolism