|Zeng, Hongmei -|
|Hatabayashi, Hidemi -|
|Nakagawa, Hiroyuki -|
|Cai, Jingjing -|
|Suzuki, Ryoya -|
|Sakuno, Emi -|
|Tanaka, Toshitsugu -|
|Ito, Yasuhiro -|
|Nakajima, Hiromitsu -|
|Yabe, Kimiko -|
Submitted to: Applied Microbiology and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 1, 2010
Publication Date: January 14, 2011
Citation: Zeng, H., Hatabayashi, H., Nakagawa, H., Cai, J., Suzuki, R., Sakuno, E., Tanaka, T., Ito, Y., Ehrlich, K., Nakajima, H., Yabe, K. 2011. Conversion of 11-hydroxy-O-methylsterigmatocystin to aflatoxin G1 in Aspergillus parasiticus. Applied Microbiology and Biotechnology. (90):635-650. Interpretive Summary: The carcinogenic compounds called aflatoxins are frequent contaminants of corn, cottonseed, peanuts and tree nuts. Much research effort has been directed to trying to prevent preharvest aflatoxin formation in these susceptible crops. Recently, we and others have found that these toxins are made by an elaborate scheme involving many proteins. In spite of the knowledge gained both from our determination of the genes required for aflatoxin production by the mold Aspergillus flavus we still have little understanding of the final steps in aflatoxin formation. The current work extends the prior knowledge of these final steps. We have proven in this paper that the final steps of aflatoxin production involve proteins residing in a membrane and non-membrane portions of the cell. One single enzyme, namely an oxidative enzyme called OrdA is responsible for multiple steps in the conversion of a precursor called HOMST to aflatoxins B1 and G1. Previous studies had only found that this enzyme was needed for formation of aflatoxin B1. The work is important because the development of specific inhibitors could involve targeting these conversion steps to prevent aflatoxin contamination.
Technical Abstract: In aflatoxin biosynthesis, aflatoxins G1 (AFG1) and B1 (AFB1) are independently produced from a common precursor, O-methylsterigmatocystin (OMST). Recently, 11-hydroxy-O-methylsterigmatocystin (HOMST) was identified as a later precursor involved in the conversion of OMST to AFB1. However, the involvement of HOMST in AFG1 formation has not been determined. We now show that HOMST is a precursor of AFG1 as well as of AFB1, and that at least two additional enzymes are required for production of AFG1 from HOMST. HOMST was prepared from OMST by incubating OrdA-expressing yeast with OMST. Feeding of Aspergillus parasiticus mutants unable to produce aflatoxins with HOMST allowed production of AFG1 as well as AFB1. Using cell-free systems, HOMST was converted to AFG1 when a microsomal fraction, cytosolic fraction from aflatoxin-producing fungi and OrdA from yeast expressing A. parasiticus OrdA were added. These results show that OrdA is required for each of at least three steps involved in conversion of OMST to AFG1 and AFB1.