title: Structure-based engineering of angucyclinone 6-ketoreductases.
abstract: Angucyclines are tetracyclic polyketides produced by Streptomyces bacteria that exhibit notable biological activities. The great diversity of angucyclinones is generated in tailoring reactions, which modify the common benz[a]anthraquinone carbon skeleton. In particular, the opposite stereochemistry of landomycins and urdamycins/gaudimycins at C-6 is generated by the short-chain alcohol dehydrogenases/reductases LanV and UrdMred/CabV, respectively. Here we present crystal structures of LanV and UrdMred in complex with NADP(+) and the product analog rabelomycin, which enabled us to identify four regions associated with the functional differentiation. The structural analysis was confirmed in chimeragenesis experiments focusing on these regions adjacent to the active site cavity, which led to reversal of the activities of LanV and CabV. The results surprisingly indicated that the conformation of the substrate and the stereochemical outcome of 6-ketoreduction appear to be intimately linked.
authors.affiliation: Department of Biochemistry, University of Turku, 20014 Turku, Finland.
authors.forename: Pekka
authors.initials: P
authors.lastname: Patrikainen
authors.affiliation: Department of Biochemistry, University of Turku, 20014 Turku, Finland.
authors.forename: Laila
authors.initials: L
authors.lastname: Niiranen
authors.affiliation: Department of Biochemistry, University of Turku, 20014 Turku, Finland.
authors.forename: Keshav
authors.initials: K
authors.lastname: Thapa
authors.affiliation: Department of Biochemistry, University of Turku, 20014 Turku, Finland.
authors.forename: Pasi
authors.initials: P
authors.lastname: Paananen
authors.affiliation: Department of Chemistry, University of Turku, 20014 Turku, Finland.
authors.forename: Petri
authors.initials: P
authors.lastname: Tähtinen
authors.affiliation: Department of Biochemistry, University of Turku, 20014 Turku, Finland.
authors.forename: Pekka
authors.initials: P
authors.lastname: Mäntsälä
authors.affiliation: Department of Biochemistry, University of Turku, 20014 Turku, Finland.
authors.forename: Jarmo
authors.initials: J
authors.lastname: Niemi
authors.affiliation: Department of Biochemistry, University of Turku, 20014 Turku, Finland. Electronic address: mikko.mk@gmail.com.
authors.forename: Mikko
authors.initials: M
authors.lastname: Metsä-Ketelä
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D018360:Crystallography, X-Ray; D016695:Glycosyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D016296:Mutagenesis; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D016415:Sequence Alignment; D013302:Streptomyces / Q000201:enzymology; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D006899:Mixed Function Oxygenases; D016695:Glycosyltransferases; C514966:LanV enzyme, Streptomyces