Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, 10029, USA.
Department of Biochemistry and Molecular Biology, School of Neurobiology, Biochemistry and Biophysics, The George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
Nat Commun. 2020 Apr 21;11(1):1916. doi: 10.1038/s41467-020-15698-8.
mHsp60-mHsp10 assists the folding of mitochondrial matrix proteins without the negative ATP binding inter-ring cooperativity of GroEL-GroES. Here we report the crystal structure of an ATP (ADP:BeF-bound) ground-state mimic double-ring mHsp60-(mHsp10) football complex, and the cryo-EM structures of the ADP-bound successor mHsp60-(mHsp10) complex, and a single-ring mHsp60-mHsp10 half-football. The structures explain the nucleotide dependence of mHsp60 ring formation, and reveal an inter-ring nucleotide symmetry consistent with the absence of negative cooperativity. In the ground-state a two-fold symmetric H-bond and a salt bridge stitch the double-rings together, whereas only the H-bond remains as the equatorial gap increases in an ADP football poised to split into half-footballs. Refolding assays demonstrate obligate single- and double-ring mHsp60 variants are active, and complementation analysis in bacteria shows the single-ring variant is as efficient as wild-type mHsp60. Our work provides a structural basis for active single- and double-ring complexes coexisting in the mHsp60-mHsp10 chaperonin reaction cycle.
mHsp60-mHsp10 有助于折叠线粒体基质蛋白，而无需 GroEL-GroES 的负 ATP 结合环间协同作用。在这里，我们报告了一个 ATP（ADP：BeF 结合）基态模拟双环 mHsp60-（mHsp10）足球复合物的晶体结构，以及 ADP 结合后继 mHsp60-（mHsp10）复合物和单环 mHsp60-mHsp10 半足球的 cryo-EM 结构。这些结构解释了 mHsp60 环形成的核苷酸依赖性，并揭示了环间核苷酸对称性与负协同作用的缺失一致。在基态中，一个二倍对称氢键和盐桥将双环缝合在一起，而当 ADP 足球准备分裂成半足球时，只有氢键仍然存在，因为赤道间隙增加。重折叠测定表明必需的单环和双环 mHsp60 变体是活跃的，并且在细菌中的互补分析表明单环变体与野生型 mHsp60 一样有效。我们的工作为 mHsp60-mHsp10 伴侣蛋白反应循环中同时存在的活性单环和双环复合物提供了结构基础。
