ATP-independent Thermoprotective Activity of Nicotiana tabacum Heat Shock Protein 70 in Escherichia coli
| Autor: | Eun Kyung Cho, Song-Ja Bae |
|---|---|
| Rok vydání: | 2007 |
| Předmět: |
Cell Survival
Nicotiana tabacum Biology medicine.disease_cause Biochemistry law.invention Sepharose Adenosine Triphosphate Affinity chromatography law Heat shock protein Tobacco Escherichia coli medicine HSP70 Heat-Shock Proteins Cloning Molecular Luciferases Molecular Biology Temperature General Medicine biology.organism_classification Molecular biology Fusion protein Recombinant Proteins Hsp70 Recombinant DNA Molecular Chaperones |
| Zdroj: | BMB Reports. 40:107-112 |
| ISSN: | 1976-6696 |
| Popis: | 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 Ni(2+) affinity chromatography and Q Sepharose Fast Flow anion exchange chromatography. The expressed fusion protein, H(6)NtHSP70-2 (hexahistidine-tagged Nicotiana tabacum heat shock protein 70-2), maintained the stability of E. coli proteins up to 90 degrees 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 degrees C) for recombinant H(6)NtHSP70-2, E. coli cells overexpressing H(6)NtHSP70-2 survived about seven times longer than those lacking H(6)NtHSP70-2. After 2 h at 50 degrees C, only the E. coli overexpressing H(6)NtHSP70-2 survived under such conditions. Our NtHSP70-2 bioassays, as well as in vitro studies, strongly suggest that HSP70 confers thermo-tolerance to E. coli. |
| Databáze: | OpenAIRE |
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