Young J C, Hartl F U
Cellular Biochemistry, Max Planck Institute for Biochemistry, Am Klopferspitz 18a, 82152 Martinsried, Germany.
EMBO J. 2000 Nov 1;19(21):5930-40. doi: 10.1093/emboj/19.21.5930.
The molecular chaperone Hsp90 binds and hydrolyses ATP, but how this ATPase activity regulates the interaction of Hsp90 with a polypeptide substrate is not yet understood. Using the glucocorticoid receptor ligand binding domain as a substrate, we show that dissociation of Hsp90 from bound polypeptide depends on the Hsp90 ATPase and is blocked by geldanamycin, a specific ATPase inhibitor. The co-chaperone p23 greatly stimulates Hsp90 substrate release with ATP, but not with the non-hydrolysable nucleotides ATPgammaS or AMP-PNP. Point mutants of Hsp90 with progressively lower ATPase rates are progressively slower in ATP-dependent substrate release but are still regulated by p23. In contrast, ATPase-inactive Hsp90 mutants release substrate poorly and show no p23 effect. These results outline an ATP-driven cycle of substrate binding and release for Hsp90 which differs from that of other ATP-driven chaperones. Conversion of the ATP state of Hsp90 to the ADP state through hydrolysis is required for efficient release of substrate polypeptide. p23 couples the ATPase activity to polypeptide dissociation and thus can function as a substrate release factor for Hsp90.
分子伴侣Hsp90能结合并水解ATP，但这种ATP酶活性如何调节Hsp90与多肽底物的相互作用尚不清楚。以糖皮质激素受体配体结合域作为底物，我们发现Hsp90从结合的多肽上解离取决于Hsp90的ATP酶活性，且会被特异性ATP酶抑制剂格尔德霉素所阻断。辅助伴侣蛋白p23能极大地促进Hsp90与ATP共同作用时底物的释放，但与不可水解的核苷酸ATPγS或AMP-PNP共同作用时则不然。ATP酶活性逐渐降低的Hsp90点突变体在依赖ATP的底物释放过程中逐渐变慢，但仍受p23的调节。相比之下，无ATP酶活性的Hsp90突变体释放底物的能力较差，且未表现出p23的作用效果。这些结果勾勒出了Hsp90由ATP驱动的底物结合与释放循环，这与其他由ATP驱动的伴侣蛋白不同。通过水解将Hsp90的ATP状态转化为ADP状态是有效释放底物多肽所必需的。p23将ATP酶活性与多肽解离相偶联，因此可作为Hsp90的底物释放因子发挥作用。
