Purification and characterization of chitinase B from moderately thermophilic bacterium Ralstonia sp. A-471.
Chitinase B was purified from a culture medium of Ralstonia sp. A-471 by precipitation with (NH4)2SO4 and column chromatography with DEAE-Toyopearl 650 M and Sephacryl S-200. The purified enzyme was homogeneous on SDS-PAGE. The molecular weight was 45,000 by SDS-PAGE. The optimum pH was 5.0 and stable pH was from 5.0 to 10.0. In the early stage of the reaction, chitinase B produced beta-anomer of (GlcNAc)2 from the substrate (GlcNAc)6, whereas (GlcNAc)4 produced almost at equilibrium, indicating that the enzyme predominantly hydrolyzes the second glycosidic linkage from the nonreducing end of (GlcNAc)6.
Ueda
Mitsuhiro
M
Laboratory of Biocycle Engineering, Department of Applied Biological Chemistry, Osaka Prefecture University, Osaka 599-8531, Japan. mueda@biochem.osakafu-u.ac.jp
Kotani
Yukiko
Y
Sutrisno
Aji
A
Nakazawa
Masami
M
Miyatake
Kazutaka
K
D002688:Chitinases / Q000302:isolation & purification* / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D006868:Hydrolysis; D043367:Ralstonia / Q000201:enzymology*; D013997:Time Factors
D002688:Chitinases