RESUMO
WYK-1 is a dipeptidyl peptidase IV inhibitor produced by Aspergillus oryzae strain AO-1. Because WYK-1 is an isoquinoline derivative consisting of three l-amino acids, we hypothesized that a nonribosomal peptide synthetase was involved in its biosynthesis. We identified 28 nonribosomal peptide synthetase genes in the sequenced genome of A. oryzae RIB40. These genes were also identified in AO-1. Among them, AO090001000009 (wykN) was specifically expressed under WYK-1-producing conditions in AO-1. Therefore, we constructed wykN gene disruptants of AO-1 after nonhomologous recombination was suppressed by RNA interference to promote homologous recombination. Our results demonstrated that the disruptants did not produce WYK-1. Furthermore, the expression patterns of 10 genes downstream of wykN were similar to the expression pattern of wykN under several conditions. Additionally, homology searches revealed that some of these genes were predicted to be involved in WYK-1 biosynthesis. Therefore, we propose that wykN and the 10 genes identified in this study constitute the WYK-1 biosynthetic gene cluster.
Assuntos
Aspergillus oryzae/enzimologia , Aspergillus oryzae/metabolismo , Dipeptidil Peptidase 4/metabolismo , Inibidores Enzimáticos/metabolismo , Isoquinolinas/metabolismo , Peptídeo Sintases/metabolismo , Aspergillus oryzae/genética , Vias Biossintéticas/genética , DNA Fúngico/química , DNA Fúngico/genética , Deleção de Genes , Dados de Sequência Molecular , Família Multigênica , Peptídeo Sintases/genética , Análise de Sequência de DNARESUMO
We identified a novel fermented substance (FKA) produced by the edible Aspergillus oryzae strain AO-1 that inhibited dipeptidyl peptidase IV (DPP-IV) (IC(50) = 3.41 mg · mL(-1)). HPLC analysis of FKA showed specific one metabolite (WYK-1), which inhibited DPP-IV (IC(50) = 6.98 µM). WYK-1 was identified as a tetrahydroxyisoquinoline derivative. Interestingly, we examined 60 strains of A. oryzae, AO-1 was the only A. oryzae strain that produces WYK-1. This is the first report that an edible A. oryzae strain produces a DPP-IV inhibitor such as WYK-1. This study also suggests that FKA has applications in the development of novel antihyperglycemic therapeutics or functional foods.
