Construction and characterization of the hetero-oligomer of the group II chaperonin from the hyperthermophilic archaeon, Thermococcus sp. strain KS-1
Construction and characterization of the hetero-oligomer of the group II chaperonin from the hyperthermophilic archaeon, Thermococcus sp. strain KS-1
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DOI：
10.1007/s00792-009-0229-3
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发表时间：
2009-02
期刊：
影响因子：
2.9
通讯作者：
M. Sahlan;Taro Kanzaki;M. Yohda
中科院分区：
文献类型：
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作者：
M. Sahlan;Taro Kanzaki;M. Yohda
超嗜热古菌Thermococcussp.菌株KS-1（T.KS-1）表达两种不同的伴侣蛋白亚基α和β，用于其蛋白质的折叠。十六聚体双环中亚基的组成随温度而变化。随着温度的升高，β亚基的含量显著增加。β亚基的同源寡聚体Cpnβ比α亚基的同源寡聚体Cpnα更耐热。由于Cpnα和Cpnβ也具有不同的蛋白折叠活性和与前折叠蛋白的相互作用，因此认为异源寡聚体根据亚基含量表现出不同的特性。然而，T.KS-1伴侣蛋白的异源寡聚体尚未研究，因为当α和β亚基同时表达时，它们形成不同组成的异源寡聚体。本研究中，我们鉴定了T.KS-1伴侣蛋白杂寡聚体Cpnαβ，它是由α和β亚基以协调方式表达并经蛋白酶消化而构建的，其中α和β以交替顺序存在。Cpnαβ保护柠檬酸合酶免于热聚集，以ATP依赖的方式促进酸变性的GFP的折叠，并表现出ATP依赖的构象变化。Cpnαβ产生的GFP复性产量与Cpnβ产生的GFP复性产量几乎相当，但低于Cpnα产生的GFP复性产量。相比之下，Cpnαβ的热稳定性与Cpnα几乎相同，但低于Cpnβ。Cpnαβ对前折叠蛋白的亲和力介于Cpnα和Cpnβ之间。
The hyperthermophilic archaeonThermococcussp. strain KS-1 (T.KS-1) expresses two different chaperonin subunits, α and β, for the folding of its proteins. The composition of the subunits in the hexadecameric double ring changes with temperature. The content of the β subunit significantly increases according to the increase in temperature. The homo-oligomer of the β subunit, Cpnβ, is more thermostable than that of the α subunit, Cpnα. Since Cpnα and Cpnβ also have different protein folding activities and interactions with prefoldin, the hetero-oligomer is thought to exhibit different characteristics according to the content of subunits. The hetero-oligomer of theT.KS-1 chaperonin has not been studied, however, because the α and β subunits form hetero-oligomers of varying compositions when they are expressed simultaneously. In this study, we characterized theT.KS-1 chaperonin hetero-oligomer, Cpnαβ, containing both α and β in the alternate order, which was constructed by the expression of α and β subunits in a coordinated fashion and protease digestion. Cpnαβ protected citrate synthase from thermal aggregation, promoted the folding of acid-denatured GFP in an ATP-dependent manner, and exhibited an ATP-dependent conformational change. The yield of refolded GFP generated by Cpnαβ was almost equivalent to that generated by Cpnβ but lower than that generated by Cpnα. In contrast, Cpnαβ exhibited almost the same level of thermal stability as Cpnα, which was lower than that of Cpnβ. The affinity of Cpnαβ to prefoldin was found to be between those of Cpnα and Cpnβ, as expected.