title: Expression, purification and preliminary X-ray diffraction analysis of a ketoreductase from a type II polyketide synthase.
abstract: Polyketide metabolites produced by bacteria and other organisms include antibiotics, anticancer and antifungal compounds. In type II polyketide synthesis, three enzymes are sufficient to form a polyketide product of the requisite chain length, although the fidelity of the first cyclization is variable. Addition of ketoreductase (KR) to this system results in the formation of a product with correct cyclization and reduction. This paper reports the cloning of the Streptomyces coelicolor actIII ORF5 gene that codes for the ketoreductase. The 261-amino-acid protein has been overexpressed with a 20-residue His tag, purified by affinity chromatography and crystallized in space group P3(2)21, with unit-cell parameters a = b = 103.9, c = 123.1 angstroms. The crystals diffract to 2.5 angstroms resolution. A complete data set has been collected and structure solution and refinement is under way.
authors.affiliation: Molecular Recognition Centre, University of Bristol, School of Medical Sciences, University Walk, Bristol BS8 1TD, England.
authors.forename: Watchrra
authors.initials: W
authors.lastname: Teartasin
authors.forename: Claire
authors.initials: C
authors.lastname: Limpkin
authors.forename: Frank
authors.initials: F
authors.lastname: Glod
authors.forename: James
authors.initials: J
authors.lastname: Spencer
authors.forename: Russell J
authors.initials: RJ
authors.lastname: Cox
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Simpson
authors.forename: John
authors.initials: J
authors.lastname: Crosby
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Crump
authors.forename: Andrea T
authors.initials: AT
authors.lastname: Hadfield
mesh_terms: D064431:3-Oxoacyl-(Acyl-Carrier-Protein) Reductase; D000429:Alcohol Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D006639:Histidine / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D016366:Open Reading Frames; D048630:Polyketide Synthases / Q000737:chemistry*; D011487:Protein Conformation; D048372:Streptomyces coelicolor / Q000235:genetics / Q000378:metabolism; D014961:X-Ray Diffraction / Q000379:methods*
chemical_list: D006639:Histidine; D048630:Polyketide Synthases; D000429:Alcohol Oxidoreductases; D064431:3-Oxoacyl-(Acyl-Carrier-Protein) Reductase