title: Structural and functional studies of a trans-acyltransferase polyketide assembly line enzyme that catalyzes stereoselective α- and β-ketoreduction.
abstract: While the cis-acyltransferase modular polyketide synthase assembly lines have largely been structurally dissected, enzymes from within the recently discovered trans-acyltransferase polyketide synthase assembly lines are just starting to be observed crystallographically. Here we examine the ketoreductase (KR) from the first polyketide synthase module of the bacillaene nonribosomal peptide synthetase/polyketide synthase at 2.35-Å resolution. This KR naturally reduces both α- and β-keto groups and is the only KR known to do so during the biosynthesis of a polyketide. The isolated KR not only reduced an N-acetylcysteamine-bound β-keto substrate to a D-β-hydroxy product, but also an N-acetylcysteamine-bound α-keto substrate to an L-α-hydroxy product. That the substrates must enter the active site from opposite directions to generate these stereochemistries suggests that the acyl-phosphopantetheine moiety is capable of accessing very different conformations despite being anchored to a serine residue of a docked acyl carrier protein. The features enabling stereocontrolled α-ketoreduction may not be extensive since a KR that naturally reduces a β-keto group within a cis-acyltransferase polyketide synthase was identified that performs a completely stereoselective reduction of the same α-keto substrate to generate the D-α-hydroxy product. A sequence analysis of trans-acyltransferase KRs reveals that a single residue, rather than a three-residue motif found in cis-acyltransferase KRs, is predictive of the orientation of the resulting β-hydroxyl group.
authors.affiliation: Institute for Cellular and Molecular Biology, The University of Texas at Austin, Texas, 78712.
authors.forename: Shawn K
authors.initials: SK
authors.lastname: Piasecki
authors.forename: Jianting
authors.initials: J
authors.lastname: Zheng
authors.forename: Abram J
authors.initials: AJ
authors.lastname: Axelrod
authors.forename: Madeline E
authors.initials: ME
authors.lastname: Detelich
authors.forename: Adrian T
authors.initials: AT
authors.lastname: Keatinge-Clay
mesh_terms: D000217:Acyltransferases / Q000737:chemistry*; D000429:Alcohol Oxidoreductases / Q000737:chemistry* / Q000648:ultrastructure*; D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000648:ultrastructure*; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010204:Pantetheine / Q000031:analogs & derivatives / Q000737:chemistry; D010453:Peptide Synthases; D011090:Polyenes; D048630:Polyketide Synthases / Q000737:chemistry*; D061065:Polyketides / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D011090:Polyenes; D061065:Polyketides; C096634:bacillaene; D010204:Pantetheine; D048630:Polyketide Synthases; D000429:Alcohol Oxidoreductases; C087192:polyketide synthase ketoreductase; D000217:Acyltransferases; D010453:Peptide Synthases; C003129:4'-phosphopantetheine
keywords: biocatalysis; crystallography; enzymology; polyketide synthase; α-ketoreduction