Laine Jennifer M, Amat Miguel, Morgan Brittany R, Royer William E, Massi Francesca
Department of Biochemistry and Molecular Pharmacology, University of Massachusetts , Worcester, Massachusetts 01605, United States.
Biochemistry. 2014 Nov 25;53(46):7199-210. doi: 10.1021/bi500591s. Epub 2014 Nov 14.
Allosteric regulation is an essential function of many proteins that control a variety of different processes such as catalysis, signal transduction, and gene regulation. Structural rearrangements have historically been considered the main means of communication between different parts of a protein. Recent studies have highlighted the importance, however, of changes in protein flexibility as an effective way to mediate allosteric communication across a protein. Scapharca dimeric hemoglobin (HbI) is the simplest possible allosteric system, with cooperative ligand binding between two identical subunits. Thermodynamic equilibrium studies of the binding of oxygen to HbI have shown that cooperativity is an entropically driven effect. The change in entropy of the system observed upon ligand binding may arise from changes in the protein, the ligand, or the water of the system. The goal of this study is to determine the contribution of the change in entropy of the protein backbone to HbI cooperative binding. Molecular dynamics simulations and nuclear magnetic resonance relaxation techniques have revealed that the fast internal motions of HbI contribute to the cooperative binding to carbon monoxide in two ways: (1) by contributing favorably to the free energy of the system and (2) by participating in the cooperative mechanism at the HbI subunit interface. The internal dynamics of the weakly cooperative HbI mutant, F97Y, were also investigated with the same methods. The changes in backbone NH dynamics observed for F97Y HbI upon ligand binding are not as large as for the wild type, in agreement with the reduced cooperativity observed for this mutant. The results of this study indicate that interface flexibility and backbone conformational entropy of HbI participate in and are important for the cooperative mechanism of carbon monoxide binding.
别构调节是许多蛋白质的一项基本功能，这些蛋白质控制着各种不同的过程，如催化、信号转导和基因调控。结构重排历来被认为是蛋白质不同部分之间通讯的主要方式。然而，最近的研究强调了蛋白质柔性变化作为介导蛋白质间别构通讯的一种有效方式的重要性。紫贻贝二聚体血红蛋白（HbI）是最简单的别构系统，两个相同亚基之间存在协同配体结合。对氧气与HbI结合的热力学平衡研究表明，协同作用是一种由熵驱动的效应。配体结合时观察到的系统熵变可能源于蛋白质、配体或系统中的水的变化。本研究的目的是确定蛋白质主链熵变对HbI协同结合的贡献。分子动力学模拟和核磁共振弛豫技术表明，HbI的快速内部运动以两种方式促进与一氧化碳的协同结合：（1）通过对系统自由能做出有利贡献；（2）通过参与HbI亚基界面处的协同机制。还使用相同方法研究了弱协同HbI突变体F97Y的内部动力学。与该突变体观察到的协同性降低一致，配体结合时F97Y HbI主链NH动力学的变化不如野生型大。本研究结果表明，HbI的界面柔性和主链构象熵参与并对一氧化碳结合的协同机制很重要。
