A strain of the yeast Saccharomyces cerevisiae coding for KHR on the chromosome secreted a toxin that kills sensitive yeasts. The transformants of multicopy vectors carrying the KHR gene could secrete 3-4-fold the killer toxin of the donor strain. This toxic substance was purified 80-fold in specific activity from the culture filtrate by gel filtration and hydrophobic column chromatography. The purified toxin gave a single protein band with molecular mass of 20 kDa on SDS-PAGE and had an isoelectric point of pH 5.3. The toxin had novel killer activity against Candida glabrata and S. cerevisiae, but did not affect bacteria, fungi, or other yeasts.
The rice koji for sake making usually shows white, but sometimes, during storage in the air it turns brown. This turn is accelerated by some wet conditions. Three kinds of precursor of the pigment in browned rice-koji were isolated in the present work by column chromatography. A main precursor was adsorbed on alumina at pH 8.5 and eluted with 0.3N acetic acid. The eluate was then treated by Dowex 50-X8 (H) column, eluted with water and 1 N HC1, and then identified as L-dopa after derived to 3,4-di-O-benzoyl-N-benzoyl-L-dopa ethyl ester. The second precursor was isolated from ethylether soluble fraction of the water eluate described above and identified as proto catechuic acid by some chemical properties. The third precursor was not isolated owing to its chemical unstableness, though it was clearly different from these two precursors. Isolation of L-dopa from the rice koji made by culture of Aspergillus and also from sake and sake cake was first carried out in the present work.
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