Yoshida T, Yohda M, Iida T, Maruyama T, Taguchi H, Yazaki K, Ohta T, Odaka M, Endo I, Kagawa Y
Biochemical Systems Laboratory, The Institute of Physical and Chemical Research (RIKEN), Hirosawa 2-1, Wako-shi, Saitama, 351-01, Japan.
J Mol Biol. 1997 Oct 31;273(3):635-45. doi: 10.1006/jmbi.1997.1337.
To elucidate the function of group II chaperonin, the gene for the chaperonin from the hyperthermophilic archaeum Thermococcus strain KS-1 was cloned and sequenced. Two distinct genes coding for chaperonin subunits, designated alpha and beta, were obtained, and their deduced amino acid sequences are highly homologous to those of group II chaperonins from other sources. The alpha and beta subunits were individually expressed in Escherichia coli. Both of the recombinant subunits assemble to constitute the homo-oligomeric double-ring complexes, which are prone to form large aggregates. The alpha aggregate is dissociated into the typical chaperonin ring complex by incubation in buffer containing 15% (v/v) methanol, while the beta aggregate cannot be dissociated. At high temperature, both of the recombinant complexes have weak ATPase activities. They are able to arrest refolding of a chemically denatured thermophilic enzyme in the absence of ATP, and refolding is resumed when ATP is supplemented. These results suggest that homo-oligomeric complexes of the archaeal chaperonin have activity.
为阐明Ⅱ型伴侣蛋白的功能，对嗜热古菌嗜热栖热菌KS-1菌株的伴侣蛋白基因进行了克隆和测序。获得了两个编码伴侣蛋白亚基的不同基因，分别命名为α和β，其推导的氨基酸序列与其他来源的Ⅱ型伴侣蛋白高度同源。α和β亚基分别在大肠杆菌中表达。两种重组亚基组装形成同寡聚双环复合物，该复合物易于形成大的聚集体。α聚集体在含有15%（v/v）甲醇的缓冲液中孵育后可解离为典型的伴侣蛋白环复合物，而β聚集体不能解离。在高温下，两种重组复合物都具有较弱的ATP酶活性。它们能够在没有ATP的情况下阻止化学变性嗜热酶的重折叠，补充ATP后重折叠恢复。这些结果表明古菌伴侣蛋白的同寡聚复合物具有活性。
