Structure, Disorder and Dynamics in Crambin Crystals at Resolution

本提案的目标是在原子分辨率上获得蛋白质及其溶剂的图像，并测试分子动力学计算如何很好地模拟结构中原子的平均位置和迁移率。这可以通过用衍射法测定蛋白质的振动和平衡结构来完成实验，也可以通过使用经验力场来完成理论。Crambin是一种疏水性植物蛋白（MW = 4720），它的独特之处在于它的衍射超过0.83 a，并且99%的衍射数据都是在这个分辨率限制下观察到的。这意味着它的结构可以不受通常的蛋白质精制的限制而确定。虽然crambin的功能尚不清楚，但它具有局部麻醉剂的作用。上述目标将通过以下具体方法使用crambin来解决：针对0.83 A， 130 K和0.945 A， 300 K x射线数据对crambin模型进行无限制的改进；300 K时中子和x射线数据的共精化；不同温度下的crambin x射线数据的细化；全晶体分子动力学模拟及与x射线结构的比较；并分析了预测蛋白质水结构的方法。这个项目提供了一个独特的机会，利用单晶x射线衍射获得蛋白质分子的实验图像，它周围的溶剂，以及它的动态运动的极其清晰的细节。所揭示的精细特征可以与目前用于预测和改进蛋白质结构的理论相检验。事实上，豆胶蛋白的晶体是非常有序的，使这些细节的测定和测试成为可能。

The goal of this proposal is to obtain a picture of a protein and its solvent at atomic resolution and to test how well the average position and mobility of atoms in the structure are modeled by molecular dynamics calculations. This can be accomplished experimentally by determining the protein's vibration and equilibrium structure using diffraction methods and theoretically by using empirical force fields. Crambin is a hydrophobic, plant protein (MW = 4720) that is unique because it diffracts beyond 0.83 A and has 99% of its diffraction data observed at this resolution limit. This means its structure can be determined free of the usual restraints of protein refinement. Although crambin's function is not yet known, it has activity as a local anesthetic. The above goal will be addressed using crambin by the following specific means: unrestrained refinement of the crambin model against the 0.83 A, 130 K and the 0.945 A, 300 K X-ray data; corefinement of neutron and X-ray data at 300 K; refinement of crambin X-ray data at multiple temperatures; full-crystal molecular dynamics simulations and comparison with the X-ray structure; and analysis of the methods to predict protein water structure. %%% This project provides a unique opportunity to obtain an experimental picture of a protein molecule, its surrounding solvent, and its dynamic movement in extremely sharp detail using single crystal X-ray diffraction. The fine features that are revealed can be tested against theories that are currently being used to predict and refine the architecture of proteins. The fact that crystals of crambin are extremely well ordered make possible the determination and testing of these details.