Corrales F J, Fersht A R
Medical Research Council Unit for Protein Function and Design, University of Cambridge, United Kingdom.
Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4509-12. doi: 10.1073/pnas.93.9.4509.
Free GroEL binds denatured proteins very tightly: it retards the folding of barnase 400-fold and catalyzes unfolding fluctuations in native barnase and its folding intermediate. GroEL undergoes an allosteric transition from its tight-binding T-state to a weaker binding R-state on the cooperative binding of nucleotides (ATP/ADP) and GroES. The preformed GroEL.GroES.nucleotide complex retards the folding of barnase by only a factor of 4, and the folding rate is much higher than the ATPase activity that releases GroES from the complex. Binding of GroES and nucleotides to a preformed GroEL.denatured-barnase complex forms an intermediately fast-folding complex. We propose the following mechanism for the molecular chaperone. Denatured proteins bind to the resting GroEL.GroES.nucleotide complex. Fast-folding proteins are ejected as native structures before ATP hydrolysis. Slow-folding proteins enter chaperoning cycles of annealing and folding after the initial ATP hydrolysis. This step causes transient release of GroES and formation of the GroEL.denatured-protein complexes with higher annealing potential. The intermediately fast-folding complex is formed on subsequent rebinding of GroES. The ATPase activity of GroEL.GroES is thus the gatekeeper that selects for initial entry of slow-folding proteins to the chaperone action and then pumps successive transitions from the faster-folding R-states to the tighter-binding/stronger annealing T-states. The molecular chaperone acts as a combination of folding cage and an annealing machine.
游离的GroEL能非常紧密地结合变性蛋白：它使巴那斯酶的折叠速度减慢400倍，并催化天然巴那斯酶及其折叠中间体的解折叠波动。在核苷酸（ATP/ADP）和GroES协同结合时，GroEL会发生从紧密结合的T态到较弱结合的R态的变构转变。预先形成的GroEL.GroES.核苷酸复合物仅使巴那斯酶的折叠速度减慢4倍，且折叠速度远高于从复合物中释放GroES的ATP酶活性。GroES和核苷酸与预先形成的GroEL.变性巴那斯酶复合物结合形成中等快速折叠的复合物。我们提出了以下分子伴侣机制。变性蛋白与静止的GroEL.GroES.核苷酸复合物结合。快速折叠的蛋白在ATP水解之前以天然结构形式被排出。缓慢折叠的蛋白在初始ATP水解后进入退火和折叠的伴侣循环。这一步导致GroES的瞬时释放，并形成具有更高退火潜力的GroEL.变性蛋白复合物。随后GroES重新结合形成中等快速折叠的复合物。因此，GroEL.GroES的ATP酶活性是一个守门人，它选择缓慢折叠的蛋白最初进入伴侣作用，然后促使其从较快折叠的R态连续转变为结合更紧密/退火更强的T态。分子伴侣起到了折叠笼和退火机器的双重作用。
