Cloning and expression of Penicillium minioluteum dextranase in Saccharomyces cerevisiae and its exploitation as a reporter in the detection of mycotoxins.
A dextranase gene from Penicillium minioluteum (strain IMI068219) has been cloned, sequenced and expressed in Saccharomyces cerevisiae via fusion of the DNA segment encoding the mature dextranase protein with alpha-factor signal sequence, and insertion into the GAL1-controlled expression vector pYES2/CT. Galactose-induced expression yielded extracellular dextranase activity of 0.63 units/ml and cell-associated dextranase activity of 0.48 units/ml, after 24 h incubation. The dextranase construct was introduced into a strain of S. cerevisiae expressing the human cytochrome P450 3A4 (CYP3A4) and the cognate reductase, which was then used to develop a microplate toxicity bioassay. Toxicity was signalled as inhibition of dextranase activity, assayed fluorimetrically. This novel bioassay was assessed using six economically significant mycotoxins.