The Role of Protein-Ligand Contacts in Allosteric Regulation of the Escherichia coli Catabolite Activator Protein.
The Role of Protein-Ligand Contacts in Allosteric Regulation of the Escherichia coli Catabolite Activator Protein.
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DOI：
10.1074/jbc.m115.669267
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发表时间：
2015-09-04
期刊：
影响因子：
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通讯作者：
Cann MJ
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文献类型：
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作者：
Townsend PD;Rodgers TL;Glover LC;Korhonen HJ;Richards SA;Colwell LJ;Pohl E;Wilson MR;Hodgson DR;McLeish TC;Cann MJ
背景：蛋白质变构可以纯粹通过改变熵来传达。结果：CAP 中 cAMP 结合强度的改变导致熵驱动变构的变化。结论：维持变构的要求限制了 CAP 中配体结合位点的进化。意义：熵驱动的过程可以限制进化中的氨基酸共变。变构是配体与蛋白质结合改变其在远处位点的活性的基本过程。实验和理论证据都表明，变构可以通过改变慢弛豫蛋白动力学而无需构象变化来传达。大肠杆菌的分解代谢激活蛋白（CAP）是分析这种熵驱动的变构的一个例子。 CAP 中的负变构发生在相同的 cAMP 结合位点之间。因此，cAMP 结合袋的变化会影响 CAP 的变构特性。在这里，我们通过粗粒度建模、等温量热法和结构分析的结合证明，降低 CAP 对 cAMP 的亲和力可以通过配体结合的熵罚增强负协同性。变体 cAMP 配体的使用表明数据不能用蛋白质突变体之间的结构异质性来解释。我们通过计算观察到，由于第二位点 CAP 突变，CAP 和 cAMP 之间相互作用强度的改变会不同程度地改变变构协同性的变化。随着 cAMP 接触位点和 CAP 上第二个位点之间的相关运动程度增加，这些位点上计算出的双突变有导致 CAP 走向不合作的趋势。 CAP 中天然存在的共变残基对不表现出这种趋势，这表明选择压力可以对 CAP 配体结合口袋的变化进行微调，而不驱动至非合作状态。一般来说，我们假设存在进化选择压力，以在发生配体结合位点进化的蛋白质中保留缓慢松弛动力学诱导的变构。
Background: Protein allostery can be communicated purely through altered entropy. Results: Altered cAMP binding strength in CAP results in changes to entropy-driven allostery. Conclusion: The requirement to maintain allostery constrains evolution of the ligand-binding site in CAP. Significance: Entropy-driven processes can constrain amino acid covariation in evolution. Allostery is a fundamental process by which ligand binding to a protein alters its activity at a distant site. Both experimental and theoretical evidence demonstrate that allostery can be communicated through altered slow relaxation protein dynamics without conformational change. The catabolite activator protein (CAP) of Escherichia coli is an exemplar for the analysis of such entropically driven allostery. Negative allostery in CAP occurs between identical cAMP binding sites. Changes to the cAMP-binding pocket can therefore impact the allosteric properties of CAP. Here we demonstrate, through a combination of coarse-grained modeling, isothermal calorimetry, and structural analysis, that decreasing the affinity of CAP for cAMP enhances negative cooperativity through an entropic penalty for ligand binding. The use of variant cAMP ligands indicates the data are not explained by structural heterogeneity between protein mutants. We observe computationally that altered interaction strength between CAP and cAMP variously modifies the change in allosteric cooperativity due to second site CAP mutations. As the degree of correlated motion between the cAMP-contacting site and a second site on CAP increases, there is a tendency for computed double mutations at these sites to drive CAP toward noncooperativity. Naturally occurring pairs of covarying residues in CAP do not display this tendency, suggesting a selection pressure to fine tune allostery on changes to the CAP ligand-binding pocket without a drive to a noncooperative state. In general, we hypothesize an evolutionary selection pressure to retain slow relaxation dynamics-induced allostery in proteins in which evolution of the ligand-binding site is occurring.