title: Double oxidation of the cyclic nonaketide dihydromonacolin L to monacolin J by a single cytochrome P450 monooxygenase, LovA.
abstract: Lovastatin, a cyclic nonaketide from Aspergillus terreus, is a hypercholesterolemic agent and a precursor to simvastatin, a semi-synthetic cholesterol-lowering drug. The biosynthesis of the lovastatin backbone (dihydromonacolin L) and the final 2-methylbutyryl decoration have been fully characterized. However, it remains unclear how two central reactions are catalyzed, namely, introduction of the 4a,5-double bond and hydroxylation at C-8. A cytochrome P450 gene, lovA, clustered with polyketide synthase lovB, has been a prime candidate for these reactions, but inability to obtain LovA recombinant enzyme has impeded detailed biochemical analyses. The synthetic codon optimization and/or N-terminal peptide replacement of lovA allowed the lovA expression in yeast (Saccharomyces cerevisiae). Both in vivo feeding and in vitro enzyme assays showed that LovA catalyzed the conversion of dihydromonacolin L acid to monacolin L acid and monacolin J acid, two proposed pathway intermediates in the biosynthesis of lovastatin. LovA was demonstrated to catalyze the regio- and stereo-specific hydroxylation of monacolin L acid to yield monacolin J acid. These results demonstrate that LovA is the single enzyme that performs both of the two elusive oxidative reactions in the lovastatin biosynthesis.
authors.affiliation: Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary T2N 1N4, Canada.
authors.forename: Jorge
authors.initials: J
authors.lastname: Barriuso
authors.forename: Don T
authors.initials: DT
authors.lastname: Nguyen
authors.forename: Jesse W-H
authors.initials: JW
authors.lastname: Li
authors.forename: Joseph N
authors.initials: JN
authors.lastname: Roberts
authors.forename: Gillian
authors.initials: G
authors.lastname: MacNevin
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Chaytor
authors.forename: Sandra L
authors.initials: SL
authors.lastname: Marcus
authors.forename: John C
authors.initials: JC
authors.lastname: Vederas
authors.forename: Dae-Kyun
authors.initials: DK
authors.lastname: Ro
mesh_terms: D001230:Aspergillus / Q000201:enzymology; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D008148:Lovastatin / Q000096:biosynthesis / Q000235:genetics; D009281:Naphthalenes / Q000737:chemistry / Q000378:metabolism*; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000201:enzymology
chemical_list: D009281:Naphthalenes; D011994:Recombinant Proteins; C045373:dihydromonacolin L; D003577:Cytochrome P-450 Enzyme System; D008148:Lovastatin; C045376:monacolin J