title: Rational Control of Polyketide Extender Units by Structure-Based Engineering of a Crotonyl-CoA Carboxylase/Reductase in Antimycin Biosynthesis.
abstract: Bioengineering of natural product biosynthesis is a powerful approach to expand the structural diversity of bioactive molecules. However, in polyketide biosynthesis, the modification of polyketide extender units, which form the carbon skeletons, has remained challenging. Herein, we report the rational control of polyketide extender units by the structure-based engineering of a crotonyl-CoA carboxylase/reductase (CCR), in the biosynthesis of antimycin. Site-directed mutagenesis of the CCR enzyme AntE, guided by the crystal structure solved at 1.5 Å resolution, expanded its substrate scope to afford indolylmethylmalonyl-CoA by the V350G mutation. The mutant A182L selectively catalyzed carboxylation over the regular reduction. Furthermore, the combinatorial biosynthesis of heterocycle- and substituted arene-bearing antimycins was achieved by an engineered Streptomyces strain bearing AntE(V350G). These findings deepen our understanding of the molecular mechanisms of the CCRs, which will serve as versatile biocatalysts for the manipulation of building blocks, and set the stage for the rational design of polyketide biosynthesis.
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan).
authors.forename: Lihan
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan).
authors.forename: Takahiro
authors.initials: T
authors.lastname: Mori
authors.affiliation: State Key Laboratory of Bio-Organic & Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Lingling road 345, Shanghai 200032 (China).
authors.forename: Qingfei
authors.initials: Q
authors.lastname: Zheng
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan).
authors.forename: Takayoshi
authors.initials: T
authors.lastname: Awakawa
authors.affiliation: State Key Laboratory of Bio-Organic & Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Lingling road 345, Shanghai 200032 (China).
authors.forename: Yan
authors.initials: Y
authors.lastname: Yan
authors.affiliation: State Key Laboratory of Bio-Organic & Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Lingling road 345, Shanghai 200032 (China).
authors.forename: Wen
authors.initials: W
authors.lastname: Liu
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan). abei@mol.f.u-tokyo.ac.jp.
authors.forename: Ikuro
authors.initials: I
authors.lastname: Abe
mesh_terms: D044944:Acyl-CoA Dehydrogenases / Q000737:chemistry* / Q000378:metabolism*; D000968:Antimycin A / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000737:chemistry; D061065:Polyketides / Q000737:chemistry*; D011487:Protein Conformation; D015202:Protein Engineering
chemical_list: D061065:Polyketides; C032456:antimycin; D000968:Antimycin A; D044944:Acyl-CoA Dehydrogenases; C100154:acyl-CoA dehydrogenase (NADP+)
keywords: biosynthesis; crotonyl-CoA carboxylase/reductase; polyketides; protein engineering