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首页> 外文期刊>Journal of Molecular Biology >Cooperative unfolding of a metastable serpin to a molten globule suggests a link between functional and folding energy landscapes
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Cooperative unfolding of a metastable serpin to a molten globule suggests a link between functional and folding energy landscapes

机译：亚稳态丝氨酸蛋白酶抑制剂向熔融小球的协同展开表明功能性和折叠能态之间的联系

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Alpha-1 antitrypsin (alpha(1),-AT) is a member of the serpin class of protease inhibitors, and folds to a metastable state rather than its thermodynamically most stable native state. Upon cleavage by a target protease, alpha(1)-AT undergoes a dramatic conformational change to a stable form, translocating the bound protease more than 70 angstrom to form an inhibitory protease-serpin complex. Numerous mutagenesis studies on serpins have demonstrated the trade-off between the stability of the metastable state on the one hand and the inhibitory efficiency on the other. Studies of the equilibrium unfolding of serpins provide insight into this connection between structural plasticity and metastability. We studied equilibrium unfolding of wild-type alpha(1)-AT using hydrogen-deuterium/exchange mass spectrometry to characterize the structure and the stability of an equilibrium intermediate that was observed in low concentrations of denaturant in earlier studies. Our results show that the intermediate observed at low concentrations of denaturant has no protection from hydrogen-deuterium exchange, indicating a lack of stable structure. Further, differential scanning calorimetry of alpha(1)-AT at low concentrations of denaturant shows no heat capacity peak during thermal denaturation, indicating that the transition from the intermediate to the unfolded state is not a cooperative first-order-like phase transition.. Our results show that the unfolding of alpha(1)-AT involves a cooperative transition to a molten globule form, followed by a non-cooperative transition to a random-coil form as more guanidine is added. Thus, the entire alpha(1)-AT molecule consists of one cooperative structural unit rather than multiple structural domains with different stabilities. Furthermore, our results together with previous mutagenesis studies suggest a possible link between an equilibrium molten globule and a functional intermediate that may be populated during the protease inhibition. (c) 2007 Elsevier Ltd. All rights reserved.
机译：Alpha-1抗胰蛋白酶（alpha（1），-AT）是serpin类蛋白酶抑制剂的成员，并且折叠成亚稳态，而不是其热力学最稳定的天然状态。在被目标蛋白酶切割后，α（1）-AT经历了急剧的构象变化，变成了稳定的形式，使结合的蛋白酶易位超过70埃，形成了抑制性蛋白酶-丝氨酸蛋白酶抑制剂复合物。关于丝氨酸蛋白酶抑制剂的大量诱变研究已经证明，一方面在亚稳态的稳定性与另一方面的抑制效率之间进行了权衡。对丝氨酸蛋白酶抑制剂平衡展开的研究提供了对结构可塑性和亚稳性之间这种联系的见解。我们使用氢-氘/交换质谱技术研究了野生型alpha（1）-AT的平衡展开，以表征早期研究中在低浓度变性剂中观察到的平衡中间体的结构和稳定性。我们的结果表明，在低浓度的变性剂中观察到的中间体对氢-氘交换没有任何保护作用，表明缺乏稳定的结构。此外，在低浓度的变性剂中，α（1）-AT的差示扫描量热法在热变性过程中没有显示出任何热容量峰值，这表明从中间状态到未折叠状态的转变不是协同的一级像相转变。我们的结果表明，α（1）-AT的展开涉及向熔融小球形式的协同转变，随后随着更多胍的加入，非协同转变为无规螺旋形式。因此，整个alpha（1）-AT分子由一个协作的结构单元而不是具有不同稳定性的多个结构域组成。此外，我们的结果与先前的诱变研究一起表明，在蛋白酶抑制过程中，平衡的熔融小球与功能性中间体之间可能存在联系。 （c）2007 Elsevier Ltd.保留所有权利。

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