Appearance of enzyme activities catalyzing conversion of sterigmatocystin to aflatoxin B1 in late-growth-phase Aspergillus parasiticus cultures

Abstract

Two activities involved in terminal pathway conversion of sterigmatocystin to aflatoxin B1 were isolated from an aflatoxin-nonproducing mutant of Aspergillus parasiticus (avn-1), and the time course of appearance of the activities in culture was determined. Subcellular fractionation of fungal mycelia resolved the two activities into a postmicrosomal activity which catalyzed conversion of sterigmatocystin to O-methylsterigmatocystin and a microsomal activity which converted O-methylsterigmatocystin to aflatoxin B1. The two activities were absent in 24-hold cells, increased to optimum levels during the stationary phase, and then declined.

Purification of a 40-kilodalton methyltransferase active in the aflatoxin biosynthetic pathway

“…Recently, versicolorin B has been demonstrated to be a precursor of versicolorin A (25,33). There is a proposed branch point in the pathway following versiconal hemiacetal acetate production leading to the different aflatoxin structural forms B1 and B2 (6,13,32,34).…”

Conversion of a new metabolite to aflatoxin B2 by Aspergillus parasiticus

“…Cultures. Cultures of A. parasiticus SRRC 2043 and SRRC 163 were maintained as described for a previous investigation (7). Single or cofermentations of the two strains were conducted by inoculating 100 ml of liquid growth medium (1) in 250-ml flasks with about 2 x 105 viable spores of each strain in 1 ml of deionized water and incubating at 28°C with shaking on a Psycrotherm incubator shaker (New Brunswick Scientific Co., Inc.) (150 rpm) for 4 days.…”

“…The AFs or OMST produced in the cultures was developed, identified, and quantitated by one-dimensional TLC by established procedures (7). HOMST from the fungal extracts was quantitated spectrophotometrically relative to an analytical HOMST standard by using the same fluorometric method as that used for OMST quantitation (5).…”

“…Three-day-old mycelia of SRRC 163 were harvested by vacuum filtration and washed extensively with low-sugar replacement medium. The mycelia (0.5 g [wet weight]) were added to each flask containing the incubation medium, and the reactions were carried out as described previously (5,7).…”

“…This was achieved by cofermentation of two A. parasiticus mutants (SRRC 2043 and SRRC 163) which are blocked at different steps in the AF biosynthetic pathway. SRRC 2043 is impaired near the terminus of the AF pathway and accumulates OMST (5) with no production of AFs, while the SRRC 163 mutant is blocked early in the AF pathway (4) but has the enzymes required for the conversion of intermediates to AFs in the late stages of the pathway (after versicolorin A synthesis) (7,13,14). * Corresponding author.…”