Viitanen P V, Lorimer G H, Seetharam R, Gupta R S, Oppenheim J, Thomas J O, Cowan N J
Central Research and Development Department, E. I. du Pont de Nemours & Co., Wilmington, Delaware 19880-0402.
J Biol Chem. 1992 Jan 15;267(2):695-8.
Chaperonins are thought to participate in the process of protein folding in bacteria and in eukaryotic mitochondria and chloroplasts. While some chaperonins are relatively well characterized, the structures of the mammalian chaperonins are unknown. We have expressed a mammalian mitochondrial chaperonin 60 in Escherichia coli and purified the recombinant protein to homogeneity. Structural and biochemical analyses of this protein establish a single toroidal structure of seven subunits, in contrast to the homologous bacterial, fungal, and plant chaperonin 60s, which have double toroidal structures comprising two layers of seven identical subjects each. The recombinant mammalian chaperonin 60, together with the mammalian chaperonin 10 (but not with bacterial chaperonin 10), facilitates the formation of catalytically active ribulose-bisphosphate carboxylase from an unfolded state in the presence of K+ and MgATP. Analysis of the partial reactions involved in this in vitro reconstitution reveals that the single toroid of chaperonin 60 can form stable complexes with both unfolded or partially folded [35S]ribulose-bisphosphate carboxylase and mitochondrial (but not bacterial) chaperonin 10 in the presence of MgATP. We conclude that the minimal functional unit of chaperonin 60 is a single hepatmeric toroid.
伴侣蛋白被认为参与细菌以及真核生物线粒体和叶绿体中的蛋白质折叠过程。虽然一些伴侣蛋白已得到较好的表征，但哺乳动物伴侣蛋白的结构尚不清楚。我们在大肠杆菌中表达了一种哺乳动物线粒体伴侣蛋白60，并将重组蛋白纯化至同质。对该蛋白的结构和生化分析确定其为七个亚基组成的单一环形结构，这与同源的细菌、真菌和植物伴侣蛋白60不同，后者具有由两层各七个相同亚基组成的双环形结构。重组的哺乳动物伴侣蛋白60与哺乳动物伴侣蛋白10（而非细菌伴侣蛋白10）一起，在K+和MgATP存在的情况下，可促使催化活性的核酮糖-1,5-二磷酸羧化酶从未折叠状态形成。对该体外重构过程中涉及的部分反应的分析表明，在MgATP存在的情况下，伴侣蛋白60的单环可与未折叠或部分折叠的[35S]核酮糖-1,5-二磷酸羧化酶以及线粒体（而非细菌）伴侣蛋白10形成稳定的复合物。我们得出结论，伴侣蛋白60的最小功能单位是单个七聚体环。
