Son Hae-Jin, Shin Eun-Jung, Nam Soo-Wan, Kim Dong-Eun, Jeon Sung-Jong
Department of Biotechnology and Bioengineering, Dong-Eui University, Busan, Korea.
FEMS Microbiol Lett. 2007 Jan;266(1):103-9. doi: 10.1111/j.1574-6968.2006.00513.x.
The gene encoding for a putative thermosome from the hyperthermophilic crenarchaeon Aeropyrum pernix K1 (ApcpnA) was cloned and the biochemical characteristics of the resulting recombinant protein were examined. The gene (accession no. APE0907) from A. pernix K1 showed some homology with other group II chaperonins from archaea. The recombinant ApcpnA protein has a molecular mass of 60 kDa, determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and exhibited ATPase activity with an optimum temperature and pH of 90 degrees C and 5.0, respectively. The ATPase activity was found to be dependent on manganese and potassium ions, but not magnesium ion. The K(m) for ATP at pH 5.0 and 90 degrees C was 10.04 (+/- 1.31) microM, and k(cat) was determined to be 2.21 (+/- 0.11) min(-1) for the ApcpnA monomer. The recombinant ApcpnA prevents thermal aggregation of bovine rhodanese and enhances the thermal stability of alcohol dehydrogenase in vitro, indicating that the protein is suitable as a molecular chaperonin in the high-temperature environment.
克隆了来自嗜热泉古菌嗜热栖热菌K1（ApcpnA）的假定热体蛋白的编码基因，并检测了所得重组蛋白的生化特性。来自嗜热栖热菌K1的该基因（登录号APE0907）与古菌的其他II组伴侣蛋白具有一些同源性。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳测定，重组ApcpnA蛋白的分子量为60 kDa，其ATP酶活性的最适温度和pH分别为90℃和5.0。发现ATP酶活性依赖于锰离子和钾离子，而不依赖于镁离子。在pH 5.0和90℃下，ApcpnA单体对ATP的K(m)为10.04（±1.31）μM，k(cat)为2.21（±0.11）min(-1)。重组ApcpnA可防止牛硫氰酸酶的热聚集，并在体外提高乙醇脱氢酶的热稳定性，表明该蛋白适合作为高温环境下的分子伴侣蛋白。
