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Applied and Environmental Microbiology, November 2003, p. 6418-6426, Vol. 69, No. 11
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.11.6418-6426.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Involvement of Two Cytosolic Enzymes and a Novel Intermediate, 5'-Oxoaverantin, in the Pathway from 5'-Hydroxyaverantin to Averufin in Aflatoxin Biosynthesis

Emi Sakuno,1,2 Kimiko Yabe,2* and Hiromitsu Nakajima1

Faculty of Agriculture, Tottori University, Koyama, Tottori 680-8553,1 National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan2

Received 21 March 2003/ Accepted 24 July 2003

During aflatoxin biosynthesis, 5'-hydroxyaverantin (HAVN) is converted to averufin (AVR). Although we had previously suggested that this occurs in one enzymatic step, we demonstrate here that this conversion is composed of two enzymatic steps by showing that the two enzyme activities in the cytosol fraction of Aspergillus parasiticus were clearly separated by Mono Q column chromatography. An enzyme, HAVN dehydrogenase, catalyzes the first reaction from HAVN to a novel intermediate, another new enzyme catalyzes the next reaction from the intermediate to AVR, and the intermediate is a novel substance, 5'-oxoaverantin (OAVN), which was determined by physicochemical methods. We also purified both of the enzymes, HAVN dehydrogenase and OAVN cyclase, from the cytosol fraction of A. parasiticus by using ammonium sulfate fractionation and successive chromatographic steps. The HAVN dehydrogenase is a homodimer composed of 28-kDa subunits, and it requires NAD, but not NADP, as a cofactor for its activity. Matrix-assisted laser desorption ionization-time of flight mass spectrometry analysis of tryptic peptides of the purified HAVN dehydrogenase revealed that this enzyme coincides with a protein deduced from the adhA gene in the aflatoxin gene cluster of A. parasiticus. Also, the OAVN cyclase enzyme is a homodimer composed of 79-kDa subunits which does not require any cofactor for its activity. Further characterizations of both enzymes were performed.

* Corresponding author. Mailing address: National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan. Phone: 81-29-838-8050. Fax: 81-29-838-8122. E-mail: yabek{at}nfri.affrc.go.jp.

Applied and Environmental Microbiology, November 2003, p. 6418-6426, Vol. 69, No. 11
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.11.6418-6426.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Hong, S.-Y., Linz, J. E. (2008). Functional Expression and Subcellular Localization of the Aflatoxin Pathway Enzyme Ver-1 Fused to Enhanced Green Fluorescent Protein. Appl. Environ. Microbiol. 74: 6385-6396 [Abstract] [Full Text]  
  • Sakuno, E., Wen, Y., Hatabayashi, H., Arai, H., Aoki, C., Yabe, K., Nakajima, H. (2005). Aspergillus parasiticus Cyclase Catalyzes Two Dehydration Steps in Aflatoxin Biosynthesis. Appl. Environ. Microbiol. 71: 2999-3006 [Abstract] [Full Text]  
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