title: Molecular insight into substrate recognition and catalysis of Baeyer-Villiger monooxygenase MtmOIV, the key frame-modifying enzyme in the biosynthesis of anticancer agent mithramycin.
abstract: Baeyer-Villiger monooxygenases (BVMOs) have been shown to play key roles for the biosynthesis of important natural products. MtmOIV, a homodimeric FAD- and NADPH-dependent BVMO, catalyzes the key frame-modifying steps of the mithramycin biosynthetic pathway, including an oxidative C-C bond cleavage, by converting its natural substrate premithramycin B into mithramycin DK, the immediate precursor of mithramycin. The drastically improved protein structure of MtmOIV along with the high-resolution structure of MtmOIV in complex with its natural substrate premithramycin B are reported here, revealing previously undetected key residues that are important for substrate recognition and catalysis. Kinetic analyses of selected mutants allowed us to probe the substrate binding pocket of MtmOIV and also to discover the putative NADPH binding site. This is the first substrate-bound structure of MtmOIV providing new insights into substrate recognition and catalysis, which paves the way for the future design of a tailored enzyme for the chemo-enzymatic preparation of novel mithramycin analogues.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, KY 40536-0596, USA.
authors.forename: Mary A
authors.initials: MA
authors.lastname: Bosserman
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, KY 40536-0596, USA.
authors.forename: Theresa
authors.initials: T
authors.lastname: Downey
authors.affiliation: Laboratory of Molecular Biology, NIDDK, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Noinaj
authors.affiliation: Laboratory of Molecular Biology, NIDDK, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Susan K
authors.initials: SK
authors.lastname: Buchanan
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, KY 40536-0596, USA.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Rohr
mesh_terms: D000970:Antineoplastic Agents / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D001665:Binding Sites; D002384:Catalysis; D018360:Crystallography, X-Ray; D006801:Humans; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000378:metabolism*; D015394:Molecular Structure; D008926:Plicamycin / Q000096:biosynthesis*; D013379:Substrate Specificity
chemical_list: D000970:Antineoplastic Agents; D006899:Mixed Function Oxygenases; D008926:Plicamycin