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Appl. Environ. Microbiol. doi:10.1128/AEM.01245-08
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Expression profiles and physiological roles of two types of molecular chaperonins from the hyperthermophilic archaeon, Thermococcus kodakaraensis

Department of Bioscience, School of Science and Technology, Kwansei-Gakuin University, 2-1 Gakuen, Sanda, Hyogo 669-1337, Japan; Nanobiotechnology Research Center, Graduate School of Science and Technology, Kwansei-Gakuin University, 2-1 Gakuen, Sanda, Hyogo 669-1337, Japan; Department of Biotechnology, College of Life Sciences, Ritsumeikan University, Kusatsu, Shiga 525-8577, Japan

^* To whom correspondence should be addressed. Email: fujiwara-s{at}kwansei.ac.jp.

	Abstract

Thermococcus kodakaraensis possesses two chaperonins, CpkA and CpkB, and their expressions are respectively induced by the temperature down-shift and up-shift of cell cultivation. The expression levels of the chaperonins were examined using specific antibodies at various cell-growth temperatures in the logarithmic and stationary phases. At 60°C, CpkA was highly expressed in both the logarithmic and stationary phases; however, CpkB was not expressed in either phase. At 85°C, CpkA and CpkB were expressed in both phases; however, the CpkA level was decreased in the stationary one. At 93°C, CpkA was expressed only in the logarithmic phase and not in the stationary phase. In contrast, CpkB was highly expressed in both phases. RT-PCR experiments showed the same growth-phase and temperature-dependent profiles as observed by immunoblot analyses, indicating that the cpkA and cpkB expressions are regulated at the mRNA level. The cpkA or cpkB gene disruptant was then constructed, and their growth profiles were monitored. The cpkA disruptant showed poor cell growth at 60°C but no significant defects at 85°C and 93°C. On the other hand, cpkB disruption led to growth defects at 93°C but no significant defects at 60°C and 85°C. These data indicate that CpkA and CpkB are necessary for cell growth at lower and higher temperatures, respectively. The logarithmic phase-dependent expression of CpkA at 93°C suggested that CpkA participates in initial cell growth in addition to lower-temperature adaptation. Promoter mapping and quantitative analyses using the Phr (Pyrococcus heat-shock regulator) gene disruptant revealed that temperature-dependent expressions were achieved in a Phr-independent manner.