Substrates: - Products: incubation of substrates with isoform AziB alone leads to a truncated single-ring product, 2-methylbenzoic acid. Presence of thioesterase AziG acting as a chain elongation and cyclization domain is required for additional two rounds of chain extension to form the expected product, 5-methyl-1-naphthoate. None of the other thioesterase domains (AziA6, AziA7, AziA8) are capable of catalyzing the reaction
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acetyl-CoA + 5 malonyl-CoA + 3 NADPH + 3 H+
5-methyl-1-naphthoate + 6 CoA + 5 CO2 + 4 H2O + 3 NADP+
Substrates: - Products: incubation of substrates with isoform AziB alone leads to a truncated single-ring product, 2-methylbenzoic acid. Presence of thioesterase AziG acting as a chain elongation and cyclization domain is required for additional two rounds of chain extension to form the expected product, 5-methyl-1-naphthoate. None of the other thioesterase domains (AziA6, AziA7, AziA8) are capable of catalyzing the reaction
constructs harboring mutant aziB, in which site specific mutations are performed for inactivating the KR and DH domains, respectively, are introduced into Streptomyces albus, yielding the recombinant strain AL1007 (to express the KR mutant AziB, G1398A within the NADPH-binding motif GxGxxG), AL1008 (to express the KR mutant AziB, Y1549F at the conserved active site), and AL1009 (to express the DH mutant AziB, H935F within the conserved motif HxxxGxxxxP). Upon HPLC-MS analysis, AL1007, AL1008, and AL1009 fail to produce 5-methyl-NPA, confirming that the reductive and dehydrating actions governed by the KR and DH domains of AziB are essential for the 5-methyl-naphthoic acid formation
heterologous expression of aziB in Streptomyces albus. Addition of aziB1 (encoding a P450 hydroxylase) and aziB2 (encoding an O-methyltransferase) to aziB in Streptomyces albus allows for the production of 3-methoxy-5-methyl-naohthoic acid as the first building block for skeleton assembly of azinomycin B
Polyketide ring expansion mediated by a thioesterase, chain elongation and cyclization domain, in azinomycin biosynthesis: characterization of AziB and AziG
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