Submitted to: Applied and Environmental Microbiology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 10/15/2005
Publication Date: 12/1/2005
Citation: Cary, J.W., Ehrlich, K., Bland, J.M., Montalbano, B.G. 2006. The aflatoxin biosynthesis cluster gene, aflX, encodes an oxidoreductase involved in conversion of versicolorin A to demethylsterigmatocystin. Applied and Environmental Microbiology. 72(2):1096-1101. Interpretive Summary: This work describes experiments that have been conducted in an effort to better understand the mechanisms by which aflatoxins are produced by fungi. Aflatoxins are toxic and carcinogenic compounds often produced by the fungi, Aspergillus flavus and A. parasiticus, during growth on crops such as corn, peanuts, cottonseed and treenuts. Because of the potential health risks, aflatoxin contamination of food and feed crops is also of great economic importance to farmers who cannot sell their crops due to strict domestic and international regulatory guidelines with regards to aflatoxin contamination. In this study, we identified the function of a gene, aflX, involved in production of aflatoxin by the fungus. As more insight is gained on the genes and enzymes involved in aflatoxin production, researchers will be able to make logical decisions as to potential targets for interruption of the synthesis of aflatoxin, thus eliminating aflatoxin contamination of food and feed crops.
Technical Abstract: Biosynthesis of the toxic and carcinogenic aflatoxins is a complicated process involving more that 27 enzymes and regulatory factors encoded by genes in a gene cluster. Previous studies found that three enzymes are required for conversion of versicolorin A (VA), to demethylsterigmatocystin (DMST), encoded by verA, ver-1, and aflY. We now show that a fourth enzyme, encoded by the previously uncharacterized gene, aflX (ordB), is also required for this conversion. A homolog of this gene, stcQ, is present in the A. nidulans sterigmatocystin (ST) biosynthesis cluster. Disruption of aflX in Aspergillus flavus gave transformants that accumulated about 4-fold more VA and 4-fold less aflatoxin than the untransformed strain. Southern and northern blot analyses confirmed that aflX was the only gene disrupted in these transformants. Feeding ST or O-methylsterigmatocystin (OMST), but not VA or earlier precursor metabolites restored normal levels of AF production. The protein encoded by aflX is predicted to have domains typical of an NADH-dependent oxidoreductase. Some domains in the protein were also similar to those in epoxide hydrolases. nThe protein encoded by aflX is similar in structure and contains some putative domains similar to the protein encoded by avfA, a gene previously shown to be involved in averufin oxidation to hydroxyversicolorone. We postulate that both proteins are involved in novel hydrolysis reactions of intermediates formed by cytochrome P450 monooxygenease-mediated epoxidation of VA and averufin.