title: Cloning and elucidation of the FR901464 gene cluster revealing a complex acyltransferase-less polyketide synthase using glycerate as starter units.
abstract: FR901464, an antitumor natural product, represents a new class of potent anticancer small molecules targeting spliceosome and inhibiting both splicing and nuclear retention of pre-mRNA. Herein we describe the biosynthetic gene cluster of FR901464, identified by degenerate primer PCR amplification of a gene encoding the 3-hydroxy-3-methylglutaryl-CoA synthase (HCS) postulated to be involved in the biosynthesis of a β-branched polyketide from Pseudomonas sp. No. 2663. This cluster consists of twenty open reading frames (ORFs) and was localized to 93-kb DNA segment, and its involvement in FR901464 biosynthesis was confirmed by gene inactivation and complementation. FR901464 is biosynthesized by a hybrid polyketide synthase (PKS)/nonribosomal peptide synthetase (NRPS), HCS, and acyltransferases (AT)-less system. The PKS/NRPS modules feature unusual domain organization including multiple domain redundancy, inactivation, and tandem. Biochemical characterization of a glyceryl transferase and an acyl carrier protein (ACP) in the start module revealed that it incorporates D-1,3-bisphosphoglycerate, which is dephosphorylated and transferred to ACP as the starter unit. Furthermore, an oxidative Baeyer-Villiger reaction followed by chain release was postulated to form a pyran moiety. On the basis of in silico analysis and genetic and biochemical evidances, a biosynthetic pathway for FR901464 was proposed, which sets the stage to further investigate the complex PKS biochemically and engineer the biosynthetic machinery for the production of novel analogues.
authors.affiliation: State Key Laboratory of Bio-organic and Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Zhang
authors.forename: Hai-Yan
authors.initials: HY
authors.lastname: He
authors.forename: Man-Cheng
authors.initials: MC
authors.lastname: Tang
authors.forename: Yu-Min
authors.initials: YM
authors.lastname: Tang
authors.forename: Qiang
authors.initials: Q
authors.lastname: Zhou
authors.forename: Gong-Li
authors.initials: GL
authors.lastname: Tang
mesh_terms: D003001:Cloning, Molecular; D005988:Glyceric Acids / Q000737:chemistry* / Q000378:metabolism; D006904:Hydroxymethylglutaryl-CoA Synthase / Q000235:genetics* / Q000378:metabolism; D008968:Molecular Conformation; D048630:Polyketide Synthases / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011714:Pyrans / Q000378:metabolism; D013141:Spiro Compounds / Q000378:metabolism
chemical_list: C104486:FR 901464; D005988:Glyceric Acids; D011714:Pyrans; D013141:Spiro Compounds; C042971:glyceric acid; D048630:Polyketide Synthases; D006904:Hydroxymethylglutaryl-CoA Synthase