title: Structural and stereochemical analysis of a modular polyketide synthase ketoreductase domain required for the generation of a cis-alkene.
abstract: The formation of an activated cis-3-cyclohexylpropenoic acid by Plm1, the first extension module of the phoslactomycin polyketide synthase, is proposed to occur through an L-3-hydroxyacyl-intermediate as a result of ketoreduction by an A-type ketoreductase (KR). Here, we demonstrate that the KR domain of Plm1 (PlmKR1) catalyzes the formation of an L-3-hydroxyacyl product. The crystal structure of PlmKR1 revealed a well-ordered active site with a nearby Trp residue characteristic of A-type KRs. Structural comparison of PlmKR1 with B-type KRs that produce D-3-hydroxyacyl intermediates revealed significant differences. The active site of cofactor-bound A-type KRs is in a catalysis-ready state, whereas cofactor-bound B-type KRs are in a precatalytic state. Furthermore, the closed lid loop in substrate-bound A-type KRs restricts active site access from all but one direction, which is proposed to control the stereochemistry of ketoreduction.
authors.affiliation: Department of Chemistry, Portland State University, Portland, OR. 97201.
authors.forename: Shilah A
authors.initials: SA
authors.lastname: Bonnett
authors.affiliation: Chemical Biology Graduate Program, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Jonathan R
authors.initials: JR
authors.lastname: Whicher
authors.affiliation: Department of Chemistry, Portland State University, Portland, OR. 97201.
authors.forename: Kancharla
authors.initials: K
authors.lastname: Papireddy
authors.affiliation: Department of Chemistry, Portland State University, Portland, OR. 97201.
authors.forename: Galina
authors.initials: G
authors.lastname: Florova
authors.affiliation: Life Sciences Institute, University of Michigan, Ann Arbor, Michigan 48109.|Department of Biological Chemistry, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Janet L
authors.initials: JL
authors.lastname: Smith
authors.affiliation: Department of Chemistry, Portland State University, Portland, OR. 97201.
authors.forename: Kevin A
authors.initials: KA
authors.lastname: Reynolds
mesh_terms: D000429:Alcohol Oxidoreductases / Q000737:chemistry / Q000378:metabolism*; D000475:Alkenes / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D055162:Biocatalysis; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D007700:Kinetics; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D013237:Stereoisomerism; D013379:Substrate Specificity
chemical_list: D000475:Alkenes; D001426:Bacterial Proteins; D000429:Alcohol Oxidoreductases; C087192:polyketide synthase ketoreductase