Thermodynamic analysis of an antagonistic folding-unfolding equilibrium between two protein domains
Thermodynamic analysis of an antagonistic folding-unfolding equilibrium between two protein domains
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DOI：
10.1016/j.jmb.2007.05.077
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发表时间：
2007-08-10
影响因子：
5.6
通讯作者：
Loh, Stewart N.
中科院分区：
文献类型：
--
作者：
Cutler, Thomas A.;Loh, Stewart N.
一个简单的模型，制定用于分析耦合折叠展开平衡存在于一类独特的分子开关蛋白。我们先前融合了两个单结构域蛋白质，芽孢杆菌RNA酶和泛素，这样存储在一个亚基的折叠结构中的自由能被用来驱动另一个亚基的展开。在这里，我们提出了一个热力学测试的机制。拮抗相互作用由耦合自由能项Δ G（x）表示。Δ G（x）是一个结构域被另一个结构域的天然结构折叠所施加的惩罚。如果Δ G（x）= 0，则两个结构域都不感知另一个结构域，它们独立地折叠和展开。如果Delta G（x）> 0，则使一个域不稳定将使另一个域稳定，反之亦然。在Δ G（x）大于任一蛋白质的内在稳定性的极限中，则在任何给定时间只有一个结构域可以折叠。我们通过测量一系列突变体的稳定性参数来估计AG（x），这些突变体使芽孢杆菌RNA酶或泛素结构域不稳定。将数据拟合到模型中，得到的Δ G（x）值类似于4 kcal mol（-1）。Δ G（x）被认为取决于用于连接两个蛋白质的接头肽的长度，以及每个结构域的固有结构可塑性。我们预测，从目前的两个和三个氨基酸残基的长度缩短接头将增加结构和热力学耦合。(C)2007爱思唯尔有限公司保留所有权利。
A simple model is formulated for analyzing the coupled folding-unfolding equilibrium present in a unique class of molecular switch proteins. We previously fused two single-domain proteins, barnase and ubiquitin, such that the free energy stored in the folded structure of one subunit is used to drive unfolding of the other. Here, we present a thermodynamic test of that mechanism. The antagonistic interaction is represented by a coupling free energy term Delta G(x). Delta G(x) is the penalty imposed on folding of one domain by the native structure of the other. If Delta G(x) = 0, then neither domain senses the other and they fold and unfold independently. If Delta G(x) > 0, then destabilizing one domain will stabilize the other, and vice versa. In the limit where Delta G(x) is greater than the intrinsic stability of either protein, then only one domain can be folded at any given time. We estimate AG(x) by measuring stability parameters for a series of mutants that destabilize either the barnase or ubiquitin domains. Fitting the data to the model leads to a Delta G(x) value of similar to 4 kcal mol(-1). Delta G(x) is proposed to depend on both the length of the linker peptides used to join the two proteins, and on the inherent structural plasticity of each domain. We predict that shortening the linkers from their current lengths of two and three amino acid residues will increase structural and thermodynamic coupling. (C) 2007 Elsevier Ltd. All rights reserved.