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KMID : 0880220110490040641
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Journal of Microbiology
2011 Volume.49 No. 4 p.641 ~ p.641
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The ATPase activity of the G2alt gene encoding an aluminium tolerance protein from Anoxybacillus gonensis G2
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Fatih Saban Beris
Lina De Smet
Jozef Van Beeumen
Hakan Karaoglu
SABRIYE CANAKCI
Ali Osman Belduz
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Abstract
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The G2ALT gene was cloned and sequenced from the thermophilic bacterium Anoxybacillus gonensis G2. The gene is 666 bp long and encodes a protein 221 amino acids in length. The gene was overexpressed in E. coli and purified to homogeneity and biochemically characterized. The enzyme has a molecular mass of 24.5 kDa and it could be classified as a member of the family of bacterial aluminium resistance proteins based on homology searches. When this fragment was expressed in E. coli, it endowed E. coli with Al tolerance to 500 μM. The purified G2ALT protein is active at a broad pH range (pH 4.0?10.0) and temperature range (25°C?80°C) with optima of 6.0 and the apparent optimal temperature of 73°C respectively. Under optimal conditions, G2ALT exhibited a low ATPase activity with Km? and Vmax? values of 10±0.55 μM and 26.81±0.13 mg Pi released/min/mg enzyme, respectively. The ATPase activity of G2ALT requires Mg2+ and Na+ ions, while Zn2+ and Al3+ stimulate the activity. Cd2+ and Ag+ reduced the activity and Li+, Cu2+, and Co2+ inhibited the activity. Known inhibitors of most ATPases, like such as β-mercaptoethanol and ouabain, also inhibited the activity of the G2ALT. These biochemical characterizations suggested that G2ALT belongs to the PP-loop ATPase superfamily and it can be responsible for aluminium tolerance in A. gonensis G2.
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KEYWORD
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aluminium tolerance gene (G2alt), aluminium tolerance protein (G2ALT), ATPase, cloning, aluminium tolerance, Anoxybacillus gonensis G2
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