Yip C M, Yip C C, Ward M D
Department of Chemical Engineering and Materials Science, University of Minnesota, Minneapolis 55455, USA.
Biochemistry. 1998 Apr 21;37(16):5439-49. doi: 10.1021/bi9722756.
Direct measurement of the forces involved in protein-protein and protein-receptor interactions can, in principle, provide insight necessary for the advancement of structural biology, molecular biology, and the development of therapeutic proteins. The protein insulin is illustrative in this respect as the mechanisms of insulin dimer dissociation and insulin-insulin receptor binding are crucial to the efficacy of insulin medications for the control of diabetes. Insulin molecules, modified with a photochemically active azido functionality on specific residues, were attached to force microscope tips and opposing mica surfaces in configurations that would either favor or disfavor dimer formation. Force curve measurements performed in buffer solution revealed the complexity of the insulin monomer-monomer interaction with multiple unbinding events occurring upon tip retraction, suggesting disruption of discrete molecular bonds at the monomer-monomer interface. Furthermore, the force curves exhibit long-range unbinding events, consistent with considerable elongation of the insulin molecule prior to dissociation. The unbinding forces observed in this study are the result of a combination of molecular disentanglement and dimer dissociation processes.
原则上，直接测量蛋白质 - 蛋白质和蛋白质 - 受体相互作用中涉及的力，可以为结构生物学、分子生物学的发展以及治疗性蛋白质的开发提供必要的见解。蛋白质胰岛素在这方面具有代表性，因为胰岛素二聚体解离和胰岛素 - 胰岛素受体结合的机制对于控制糖尿病的胰岛素药物疗效至关重要。在特定残基上用具有光化学活性的叠氮基官能团修饰的胰岛素分子，以有利于或不利于二聚体形成的构型附着在力显微镜尖端和相对的云母表面上。在缓冲溶液中进行的力曲线测量揭示了胰岛素单体 - 单体相互作用的复杂性，在尖端回缩时发生多次解离事件，这表明在单体 - 单体界面处离散分子键的破坏。此外，力曲线显示出长程解离事件，这与胰岛素分子在解离前的显著伸长一致。本研究中观察到的解离力是分子解缠和二聚体解离过程共同作用的结果。
