PURIFICATION OF 3 CATALASE ISOZYMES FROM FACULTATIVELY ALKALIPHILIC BACILLUS-FIRMUS OF4

被引:22
作者
HICKS, DB
机构
[1] Department of Biochemistry, Mount Sinai School of Medicine, CUNY, NT, 10029, One Gustave L. Leuy Place, New York
来源
BIOCHIMICA ET BIOPHYSICA ACTA-BIOENERGETICS | 1995年 / 1229卷 / 03期
关键词
CATALASE; HYDROGEN PEROXIDE; ALKALIPHILE; PH;
D O I
10.1016/0005-2728(95)00016-C
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Cell extracts of facultatively alkaliphilic B. firmus OF4 were assayed for catalase activity and their catalase isozyme content was analyzed on native polyacrylamide gels stained for catalase activity. pH-10.5-grown cells had about twice the specific catalase activity of pH-7.5-grown cells. The higher activity, however, did not confer resistance to exogenous hydrogen peroxide challenge relative to pH-7.5-grown cells and, in fact, the pH-10.5-grown cells were much more sensitive to the challenge. Electrophoresis resolved three catalase isozymes in cell extracts. The isozymes, labeled I-III in order of decreasing electrophoretic mobility, were purified and their Nterminal amino acid sequences were obtained. Isozyme III corresponded to the product of a cloned gene fragment that had been shown to possess substantial sequence similarity to the KatE (HP-II) catalase of E, coli (Quirk, P.G., Krulwich, T.A. and Hicks, D.B. (1993) Biophys. J. 64, 164A) and which had similar biochemical properties to HP-II, i.e., it was a chlorin-containing enzyme expressed only in stationary phase. Isozyme II, a protoheme enzyme, was responsible for the higher activity of alkaline-grown cells and was induced in cells treated with hydrogen peroxide or ascorbate. It showed sequence similarity to katA of Bacillus subtilis (Bol, D, and Yasbin, R. (1991) Gene 109, 31-37). Isozyme I was the only isozyme that exhibited detectable levels of peroxidase activity in addition to catalase activity, resembling a catalase enzyme purified from a different alkaliphile, Bacillus YN-2000 (Yumoto, I., Fukumori, Y. and Yamanaka, T. (1990) J. Biochem. 108, 583-587), to which it showed some sequence similarity.
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页码:347 / 355
页数:9
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