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The prosegment catalyzes pepsin folding to a kinetically trapped native state

机译:突变催化胃蛋白酶折叠到动力学上捕获的天然状态

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Investigations of irreversible protein unfolding often assume that alterations to the unfolded state, rather than the nature of the native state itself, are the cause of the irreversibility. However, the present study describes a less common explanation for the irreversible denaturation of pepsin, a zymogen-derived aspartic peptidase. The presence of a large folding barrier combined with the thermodynamically metastable nature of the native state, the formation of which depends on a separate prosegment (PS) domain, is the source of the irreversibility. Pepsin is unable to refold to the native state upon return from denaturing conditions due to a large folding barrier (24.6 kcal/mol) and instead forms a thermodynamically stable, yet inactive, refolded state. The native state is kinetically stabilized by an unfolding activation energy of 24.5 kcal/mol, comparable to the folding barrier, indicating that native pepsin exists as a thermodynamically metastable state. However, in the presence of the PS, the native state becomes thermodynamically stable, and the PS catalyzes pepsin folding by stabilizing the folding transition state by 14.7 kcal/mol. Once folded, the PS is removed, and the native conformation exists as a kinetically trapped state. Thus, while PS-guided folding is thermodynamically driven, without the PS the pepsin energy landscape is dominated by kinetic barriers rather than by free energy differences between native and denatured states. As pepsin is the archetype of a broad class of aspartic peptidases of similar structure and function, and many require their PS for correct folding, these results suggest that the occurrence of native states optimized for kinetic rather than thermodynamic stability may be a common feature of protein design.
机译:不可逆蛋白展开的调查通常认为,改变对展开状态而不是本地国家本身的性质,是不可逆转性的原因。然而,本研究描述了对胃蛋白酶的不可逆变性的常见解释,一种百分比衍生的天冬氨酸肽酶。与天然状态的热力学亚稳态性质相结合的大折叠屏障的存在,其形成取决于单独的突变域(PS)域,是不可逆性的来源。由于大折叠屏障(24.6千卡/摩尔)而从变性条件返回,百事可乐无法重新折叠到原生状态,而是形成热力学稳定但又无效的重折叠状态。本地状态通过24.5kcal / mol的展开活化能量,与折叠屏障相当,表明天然胃蛋白酶作为热力学上稳定状态。然而,在PS存在下,天然状态变得热力学稳定,并且PS通过将折叠过渡状态稳定14.7kcal / mol来催化胃蛋白酶折叠。一旦折叠,PS被移除,并且本地构象作为动力学捕获状态存在。因此,虽然热力学驱动的PS引导折叠,但没有PS,百事可乐能量景观由动力学屏障主导而不是天然和变性状态之间的自由能差。作为百事可乐是一类相似的结构和功能的广泛天冬氨酸肽酶的原型,许多需要它们的PS进行正确的折叠,这些结果表明,对动力学而不是热力学稳定性优化的天然状态可能是蛋白质的常见特征设计。

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