Enzymatic conversion of norsolorinic acid to averufin in aflatoxin biosynthesis

Abstract

Hydroxyaverantin (HAVN) was isolated from a mold, Emericella heterothallica IFO 30842. Aspergillus parasiticus NIAH-26, a UV-irradiated mutant of A. parasiticus SYS-4, produced neither aflatoxins nor precursors in yeast extract-sucrose (YES) medium. When the postmicrosome (cytosol) fraction of NIAH-26, which had been prepared from the culture in YES medium, was incubated with norsolorinic acid (NA) in the presence of NADH or NADPH, averantin (AVN) was produced. The reverse reaction from AVN to NA was promoted by the addition of NAD or NADP (dehydrogenase reaction). When the microsome fraction of NIAH-26 was incubated with AVN, HAVN was produced in the presence of NADPH (monooxygenase reaction). HAVN was, furthermore, oxidized to averufin (AVR) by the cytosol fraction of NIAH-26 in the presence of NAD or NADP (dehydrogenase reaction). In the feeding experiments with A. parasiticus NIAH-26, aflatoxins were produced from AVN, HAVN, NA, and AVR but not from averufanin or averythrin. These results indicate that the reaction sequence NA<-*AVN-*HAVN-*AVR is involved in the biosynthetic pathway of aflatoxins. The enzyme activities described here were dependent on the culture medium, and no enzyme activities were observed in the nonaflatoxigenic strain A. oryzae SYS-2 (IFO 4251).

Purification and Gene Cloning of a Dehydrogenase fromLactobacillus brevisThat Catalyzes a Reaction Involved in Aflatoxin Biosynthesis

“…HAVN was isolated from myceria of Emericella heterothallica IFO 30842. 18) VA and nidurufin were prepared from mycelia of A. versicolor (Vuillemin) Tiraboschi. 19,20) Averantin was prepared from mycelia of the averantin-accumulating mutant A. parasiticus NIAH 204.…”

“…The concentrations of the metabolites were determined from UV absorption coefficients ( max ): for HAVN, 7,100 (466 nm), and for AVR, 10,500 (454 nm). 18) Other dehydrogenases. Alcohol dehydrogenase from L. kefir (Fluka, Tokyo) and glucose dehydrogenase (Wako, Osaka, Japan) were used in this study.…”

“…The spot having the same Rf value as AVR in lanes 1 and 4 was averufanin, converted spontaneously from HAVN. 18,20) a Cell extract was prepared from 25 g wet weight of cells harvested from 5 liters of culture medium. b The activity and protein concentration of cell extract was determined after removal of Triton X-100 using a spin column with Bio Beads SM-2.…”

“…This suggests that the reaction from HAVN to OAVN precedes the stream for aflatoxin formation. 18,38) Although both enzymes, Lac-HAVN dehydrogenase and Aspergillus HAVN dehydrogenase, catalyzed the same reaction from HAVN to OAVN, their properties were different. The native molecular masses of the Lac-HAVN dehydrogenase and A. parasiticus HAVN dehydrogenase were quite different: 100 and 60 kDa respectively, whereas the molecular masses deduced from their amino acid sequences were similar, 26 and 28 kDa respectively.…”

A red pigment synthesized by an Aspergillus parasiticus mutant as a possible new intermediate in the aflatoxin biosynthetic pathway

“…(Townsend et al 1988), methoxyaverantin (Yabe et al 1988) andversicolorin B (McGuire et al 1989). Several of the enzymes catalysing the biosynthetic steps in AFB, formation have also been isolated Dutton 1988;Bhatnagar et al 1989;Chuturgoon et al 1990;Anderson et al 1990;Yabe et al 1991a;Yabe et al 1991b).…”