A filamentous fungus belonging to Penicillium purpurescens was isolated as a strain grown well in methanol medium. The strain produced an alcohol oxidase (ACID), which oxidizes short-chain primary alcohols and ethylene glycol, by incubation with a medium containing not only methanol, but also ethanol, ethylene glycol, glycerol or glucose. This ACID belonged to the same group as AOD from methylotrophic yeast, but some properties were different from AODs from methylotrophic yeast and other fungi. Thus, this enzyme consisted of four identical subunits of 66 kDa, while AODs from methylotrophic yeasts were of eight subunits. The enzyme activity was enhanced to two-fold by incubation at pH 6.0 and 40 degrees C for 60 min, whereas the K. values for ethanol and ethylene glycol did not change. (C) 2008, The Society for Biotechnology, Japan. All rights reserved.