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KMID : 1094720070120050502
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Biotechnology and Bioprocess Engineering
2007 Volume.12 No. 5 p.502 ~ p.507
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Enhanced tolerance against freezing stress in Escherichia coli cells expressing an algal cyclophilin gene
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Cho Eun-Kyung
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Abstract
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Members of the cyclophilin (Cyp) family are known to function as co-chaperones, interacting with chaperones such as heat shock protein 90, and perform important roles in protein folding under high temperature stress. In addition, they have been isolated from a wide range of organisms. However, there have been no reports on the functions of algal Cyps under other stress conditions. To study the functions of the cDNAGjCyp-1 isolated from the red alga (Griffithsia japonica), a recombinant GjCyp-1 containing a hexahistidine tag at the amino-terminus was constructed and expressed inEscherichia coli. Most of the gene product expressed inE. coli was organized as aggregate insoluble particles known as inclusion bodies. Thus, the optimal time, temperature, and concentration ofl(+)-arabinose for expressing the soluble and nonaggregated form of GjCyp-1 inE. coli were examined. The results indicate that the induction of Cyp, at 0.2%l(+)-arabinose for 2 h at 25°C, had a marked effect on the yield of the soluble and active form of the co-chaperone as PPlase. An expressed fusion protein, H6GjCyp-1, maintained the stability ofE. coli proteins up to-75°C. In a functional bioassay of the recombinant H6GjCyp-1, the viability ofE. coli cells overexpressing H6GjCyp-1 was compared to that of cells not expressing H6GjCyp-1 at ?75°C. For all the cycles of a freeze/thaw treatment, a significant increase in viability was observed in theE. coli cells overexpressing H6GjCyp-1. The results of the GjCyp-1 bioassays, as well asin vitro studies, strongly suggest that the algal Cyp confers freeze tolerance toE. coli.
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KEYWORD
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Griffithsia japonica, cyclophilin, Escherichia coli, freezing tolerance, in vivo and in vitrofunction
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