Synthesis and Study of Minimal Protein Folding Models: Molecular Torsion Balance

描述（由申请人提供）：测试和改进我们对影响生物现象（包括蛋白质折叠、蛋白质识别、药物受体结合和酶催化等事件）的分子力的理解，需要在控制良好的实验中进行精确的观察。我们开发了一种工具——“分子扭转平衡”——用于测量两种分离良好的热力学状态的相对能量，它们只是在构象上有所不同。这些精密的分子工具可以可靠地测量小到0.05千卡/摩尔的构象的微分能量效应。我们建议在水系统中使用这些分子，并获得与五个重要结合基序相关的定量数据，这些基序是蛋白质稳定性理论的核心要素。子项目旨在提供与理解生物识别（折叠和结合）直接相关的数据，并用于测试当前的生物计算方法和理论。我们将研究：1)盐桥对构象稳定性的影响。我们的实验将提供生物分子中最常见的溶剂暴露盐桥强度的定量比较，以及离子强度和温度对这些效应的影响。2)疏水结合及非极性表面对水介导构象稳定性的影响。Lum-Chandler-Weeks （LCW）疏水性理论对疏水性如何随烷基表面尺寸的变化做出了有趣的预测。我们试图验证这一预测并量化差异。3)卤素和“卤素键”对水中构象稳定性的影响。据报道，将卤素引入药物分子会改变结合位点的亲和力。邻基对疏水相互作用的影响。附近的官能团如何影响疏水表面的“粘性”？对水的结构是否有影响，可以改变在疏水-水界面的多余自由能？5) β转模拟分子扭平衡。一个中等风险-高影响子项目将允许直接比较反平行方向成对氨基酸相互作用的能量和氨基酸变化对短链稳定性的影响。在这些实验研究中获得的知识将可用于测试当前的生物识别计算方法和理论，并确定设计生物活性剂的指导原则。

DESCRIPTION (provided by applicant): Testing and improvement in our understanding of the molecular forces that influence biological phenomena (events including protein folding, protein-protein recognition, drug-receptor binding, and enzyme catalysis) require precise observations in well-controlled experiments. We developed a tool - the 'molecular torsion balance' - for measuring the relative energies of two well separated thermodynamic states which differ only in their conformation. These precision molecular tools can reliably measure differential energy effects on conformation as small as 0.05 kcal/mol. We propose to use these molecules in aqueous systems and to acquire quantitative data relevant to five important binding motifs that are central elements in theories of protein stability. The subprojects are designed to provide data directly relevant to understanding biological recognition (folding and binding) and useful for testing current biocomputational methods and theories. We will investigate: 1) The effects of salt bridges on conformational stability. Our experiments will provide quantitative comparisons of the strength of solvent-exposed salt bridges most common in biological molecules, and on the effects of ionic strength and temperature on these effects. 2) Hydrophobic binding and the effect of non-polar surfaces on water-mediated conformational stability. The Lum-Chandler-Weeks (LCW) theory of hydrophobicity makes intriguing predictions on how hydrophobicity changes with the sizes of alkyl surfaces. We seek to verify this prediction and quantify the differences. 3) The effects of halogens and the 'halogen bond' on conformation stability in water. The introduction of halogens into drug molecules is reported to change binding site affinities. 4) Neighboring group effects on hydrophobic interactions. How do nearby functional groups influence the 'stickiness' of hydrophobic surfaces? Is there an effect on the structure of water that can change the excess free energy at hydrophobic-water interfaces? 5) A beta-turn mimetic molecular torsion balance. A moderate risk -high impact subproject will allow direct comparisons of the energy of pair-wise amino acid interactions in anti-parallel orientation and the effects of amino acid changes on short beta-strand stability. Knowledge gained in these experimental studies will be available for testing current computational methods and theories of biological recognition and in identifying guiding principles for design of biologically active agents.