Endo-beta-Glucanase from Acetobacter xylinum: Purification andCharacterization
A cellulose-producing acetic acid bacterium,Acetobacter xylinum KU-1, abundantly produces an extracellularendo-beta-glucanase (EC 3.2.1.4) in the culture broth. The enzyme was purifiedto homogeneity by DEAE- and CM- Toyopearl 650M ion-exchange chromatography,Butyl-Toyopearl 650M hydrophobic chromatography, and Toyopearl HW-50 gelfiltration. The purified enzyme showed the maximum activity at pH 5 and50&deg;C: it was stable up to 50&deg;C at pH 5, activated by Co2+, andcompetitively inhibited by Hg2+; the apparentKi was 7 &mgr;M. The molecular weight of the enzyme wasdetermined to be about 39,000 by sodium dodesyl sulfate/polyacrylamide gelelectrophoresis, and about 41,000 by Toyopearl HW-50 gel filtration; theenzyme is monomeric. The enzyme hydrolyzed carboxymethylcellulose with anapparent Km of 30 mg/ml and Vmax of 1.2&mgr;M/min. It hydrolyzed cellohexaose to cellobiose, cellotriose andcellotetraose, and also cellopentaose to cellobiose and cellotriose, but didnot act on cellobiose, cellotriose, or cellotetraose.
Oikawa
T
Department of Biotechnology, Faculty of Engineering, Kansai University, 3-3-35, Yamate-cho, Suita-shi, Osaka 564, Japan
Kamatani
T
Kaimura
T
Ameyama
M
Soda
K