Discovery of a two-component monooxygenase SnoaW/SnoaL2 involved in nogalamycin biosynthesis.
Nogalamycin is an anthracycline polyketide antibiotic that contains two deoxysugars, at positions C-1 and C-7. Previous biosynthetic studies conducted in vivo affiliated snoaL2 with an unusual C-1 hydroxylation reaction, but in vitro activity was not established. Here, we demonstrate that inactivation of either snoaL2 or snoaW resulted in accumulation of two nonhydroxylated metabolites, nogalamycinone and a novel anthracycline 3',4'-demethoxy-nogalose-nogalamycinone. The C-1 hydroxylation activity was successfully reconstructed in vitro in the presence of the two enzymes, NAD(P)H and the substrates. Based on relative reaction efficiencies, 3',4'-demethoxy-nogalose-nogalamycinone was identified as the likely natural substrate. A biosynthetic model was established where the atypical short-chain alcohol dehydrogenase SnoaW reduces the anthraquinone to a dihydroquinone using NADPH, which enables activation of oxygen and formation of a hydroperoxy intermediate. Finally, protonation of the intermediate by SnoaL2 yields the 1-hydroxylated product.
Department of Biochemistry and Food Chemistry, University of Turku, Turku FIN-20014, Finland.
Vilja
V
Siitonen
Bastian
B
Blauenburg
Pauli
P
Kallio
Pekka
P
Mäntsälä
Mikko
M
Metsä-Ketelä
D000903:Antibiotics, Antineoplastic / Q000737:chemistry / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006861:Hydrogen Peroxide / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D009621:Nogalamycin / Q000737:chemistry / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D013302:Streptomyces / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
D000903:Antibiotics, Antineoplastic; D006861:Hydrogen Peroxide; D006899:Mixed Function Oxygenases; D009621:Nogalamycin; D010100:Oxygen