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KMID : 0380220070400010107
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Journal of Biochemistry and Molecular Biology
2007 Volume.40 No. 1 p.107 ~ p.112
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ATP-independent Thermoprotective Activity of Nicotiana tabacum Heat Shock Protein 70 in Escherichia coli
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Cho Eun-Kyung
Bae Song-Ja
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Abstract
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To study the functioning of HSP70 in Escherichia coli, we selected NtHSP70-2 (AY372070) from among three genomic clones isolated in Nicotiana tabacum. Recombinant NtHSP70-2, containing a hexahistidine tag at the amino-terminus, was constructed, expressed in E. coli, and purified by Ni2+ affinity chromatography and Q Sepharose Fast Flow anion exchange chromatography. The expressed fusion protein, H6NtHSP70-2 (hexahistidine-tagged Nicotiana tabacum heat shock protein 70-2), maintained the stability of E. coli proteins up to 90°C. Measuring the light scattering of luciferase (luc) revealed that NtHSP70-2 prevents the aggregation of luc without ATP during high-temperature stress. In a functional bioassay (1 h at 50°C) for recombinant H6NtHSP70-2, E. coli cells overexpressing H6NtHSP70-2 survived about seven times longer than those lacking H6NtHSP70-2. After 2 h at 50°C, only the E. coli overexpressing H6NtHSP70-2 survived under such conditions. Our NtHSP70-2 bioassays, as well as in vitro studies, strongly suggest that HSP70 confers thermo-tolerance to E. coli.
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KEYWORD
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Escherichia coli, Heat shock protein 70 (HSP70), in vivo and in vitro Functioning, Nicotiana tabacum, Thermotolerance
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