Energetic and Structural Effects in Protein-Ligand Interactions

在有机和大分子项目的支持下，斯蒂芬·马丁教授认识到了分子识别中的一个关键挑战，即了解如何设计与蛋白质紧密结合的小分子。为此，了解在配体中引入特定的结构变化如何影响修饰配体的相对结合亲和力是至关重要的。这项任务变得更加复杂，因为焓/熵补偿总是伴随着这样的结构变化。该小组最近发现，在小分子中引入构象约束并不一定会导致更有利的结合熵，这一发现与配体前组织的假定效应相反。因此，他们提出，配体结构的变化将对蛋白质的灵活性和动力学产生不同的影响，这将表现为补偿结合热和结合熵的变化，这些变化可能会增强、减弱或超过对小分子进行结构修改的预期效果。为了验证这一假说，我们采用了多学科的方法来研究在蛋白质-配体相互作用中，配体结构的选择性变异是如何影响能量、结构和动力学的。具体地说，他们将制备受约束的、灵活的磷酸酪氨酸衍生配体，这些配体将与Grb2SH2结构域结合。他们还将制备具有不同极性和非极性表面积比率的类似物，以探测疏水效应。然后，他们将使用等温滴定量热法来确定不同配体与Grb2SH2结构域的结合。选定的配合物将通过X射线结晶学进行表征，以研究配体与结构域之间的详细相互作用。对实验数据的分析将使人们能够将配体结构的变化与能量、结构和蛋白质柔性的变化联系起来，并探索蛋白质-配体相互作用中的焓/熵补偿的来源。拟议活动的更广泛影响是，参与的研究生和本科生，特别是本科生女性和拉丁裔，将在一个生物分子识别前沿的跨学科项目中获得知识和学习实验技术。这项培训将使他们能够在有机化学、分子生物学和生物物理学的关键界面上从事职业生涯。在本科生课堂和当地一所高中展示这些发现，将有助于刺激更多的学生追求科学职业。此外，这些研究的结果将有助于形成一个核心知识，将加强我们对蛋白质-配体相互作用的理解，并促进基于结构的小分子的设计，这些小分子与蛋白质靶标具有高亲和力。由此得出的见解将使药物化学家更好地优化配体结合亲和力，因为他们将铅转化为治疗疾病的选择性和有效的治疗剂。

With the support of an award from the Organic and Macromolecular Program, Professor Stephen Martin has recognized a critical challenge in molecular recognition which is understanding how to design small molecules that bind tightly to proteins. Toward this end, understanding how introducing specific structural changes into a ligand affects the relative binding affinity of the modified ligand is of paramount importance. The task is rendered more complicated because enthalpy/entropy compensation invariably attends such structural variations. The group recently discovered that introducing conformational constraints in small molecules does not necessarily lead to more favorable binding entropies, a finding that is opposite to the putative effects of ligand preorganization. They thus suggest that variations in ligand structure will have differential consequences upon protein flexibility and dynamics that will be manifested in compensating changes in binding enthalpies and entropies that may enhance, attenuate, or override the anticipated effects of making structural modifications to a small molecule. In order to evaluate this hypothesis, a multidisciplinary approach has been adopted to investigate how selected variations in ligand structure affect energetics, structure and dynamics in protein-ligand interactions. Specifically, they will prepare constrained and flexible phosphotyrosine-derived ligands that will bind to the Grb2 SH2 domain. They will also prepare analogs having varying ratios of polar and nonpolar surface areas to probe hydrophobic effects. They will then use isothermal titration calorimetry to determine binding of various ligands to the Grb2 SH2 domain. Selected complexes will be characterized by x-ray crystallography to study the detailed interactions between the ligands and the domain. Analysis of the experimental data will enable use to correlate variations in ligand structure with changes in energetics, structure and protein flexibility and to probe the origin of enthalpy/entropy compensation in protein-ligand interactions. The broader impact of the proposed activity is that participating graduate and undergraduate students, especially undergraduate women and Latinos, will gain knowledge and learn experimental techniques in an interdisciplinary project at the forefront of biomolecular recognition. This training will enable them to pursue careers at the critical interface of organic chemistry, molecular biology and biophysics. Presenting the findings in the undergraduate classroom and a local high school will help stimulate even more students to pursue careers in science. Additionally, the results of these investigations will contribute to developing a core of knowledge that will enhance our understanding of protein-ligand interactions and facilitate the structure-based design of small molecules having high affinities for protein targets. Insights thus derived will better enable medicinal chemists to optimize ligand binding affinities as they transform leads into selective and potent therapeutic agents to treat diseases.