Viitanen P V, Donaldson G K, Lorimer G H, Lubben T H, Gatenby A A
Central Research and Development Department, E.I. du Pont de Nemours and Company, Wilmington, Delaware 19880-0402.
Biochemistry. 1991 Oct 8;30(40):9716-23. doi: 10.1021/bi00104a021.
The spontaneous refolding of chemically denatured dihydrofolate reductase (DHFR) is completely arrested by chaperonin 60 (GroEL). This inhibition presumably results from the formation of a stable complex between chaperonin 60 and one or more intermediates in the folding pathway. While sequestered on chaperonin 60, DHFR is considerably more sensitive to proteolysis, suggesting a nonnative structure. Bound DHFR can be released from chaperonin 60 with ATP, and although chaperonin 10 (GroES) is not obligatory, it does potentiate the maximum effect of ATP. Hydrolysis of ATP is also not required for DHFR release since certain nonhydrolyzable analogues are capable of partial discharge. "Native" DHFR can also form a stable complex with chaperonin 60. However, in this case, complex formation is not instantaneous and can be prevented by the presence of DHFR substrates. This suggests that native DHFR exists in equilibrium with at least one conformer which is recognizable by chaperonin 60. Binding studies with 35S-labeled DHFR support these conclusions and further demonstrate that DHFR competes for a common saturable site with another protein (ribulose-1,5-bisphosphate carboxylase) known to interact with chaperonin 60.
化学变性的二氢叶酸还原酶（DHFR）的自发重折叠被伴侣蛋白60（GroEL）完全抑制。这种抑制作用大概是由于伴侣蛋白60与折叠途径中的一种或多种中间体形成了稳定的复合物。当被隔离在伴侣蛋白60上时，DHFR对蛋白水解的敏感性大大增加，这表明其结构为非天然结构。结合的DHFR可以通过ATP从伴侣蛋白60上释放出来，虽然伴侣蛋白10（GroES）不是必需的，但它确实能增强ATP的最大效应。由于某些不可水解的类似物能够部分释放DHFR，因此DHFR的释放也不需要ATP水解。“天然”DHFR也能与伴侣蛋白60形成稳定的复合物。然而，在这种情况下，复合物的形成不是瞬间完成的，并且可以通过DHFR底物的存在来阻止。这表明天然DHFR与至少一种能被伴侣蛋白60识别的构象异构体处于平衡状态。用35S标记的DHFR进行的结合研究支持了这些结论，并进一步证明DHFR与另一种已知与伴侣蛋白60相互作用的蛋白质（核酮糖-1,5-二磷酸羧化酶）竞争一个共同的饱和位点。
