RIGID BODY REFINEMENT BY USING ESR RESTRAINTS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 蛋白质复合体的研究不仅对于理解调控途径，而且对于药物设计都是至关重要的。核磁共振或X射线结晶学在预测蛋白质复合体结构方面的范围仍然有限，因此涉及两个蛋白质对接的计算程序变得越来越重要。传统上，两个蛋白质的对接问题要么基于几何学，其中蛋白质框架以数学模型的形式表示，要么基于能量学，其中蛋白质的行为像刚性、半柔性或柔性的身体。后一种方法的主要新兴趋势之一是将实验信息包括在对接计算中。为了这个目的，这个子项目处理在ESR测量的距离约束下由两个蛋白质形成的复合体的刚体精化。精炼是通过使用称为晶体和核磁共振系统(CNS)的软件包进行的。该软件包传统上用于使用结晶学或核磁共振数据作为输入来确定大分子结构。为了根据ESR距离执行刚体精化，有必要在程序文件中进行修改。这些距离以类似于NOE距离的方式合并，NOE距离以表格的形式存在，并且每个距离(D)被分配负误差(Dminus)和正误差(Dplus)，从而考虑了自旋标签的灵活性。然后使用共轭梯度法来最小化整个系统的能量。除了取决于ESR测量距离的准确性外，精化还依赖于与每个距离相关的误差分配。目前，该程序已经在趋化蛋白Chew和Delta289的P5结构域之间形成的复合体上进行了测试，在它们之间已经测量了几个ESR距离。刚体细化给出了两种蛋白质的构象，这两种蛋白质的构象接近于复合体的晶体结构。重要的是，在没有任何其他支持数据的情况下，这两种蛋白质之间只需要12个ESR距离就可以很好地近似它们的复杂结构。然而，该程序需要在更多这样的系统上进行测试，并且需要设计一种系统的方法来分配与每个ESR距离相关的误差。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The study of protein complexes is crucial not only for the purpose of understanding regulatory pathways, but also for drug design. The scope of NMR or X-ray crystallography in predicting the structure of a protein complex is still limited, hence computational procedures involving docking of two proteins have gained importance. Traditionally, the problem of docking of two proteins is either based on geometry in which the protein frame is represented in the form of a mathematical model or energetics, in which the protein acts like a rigid, semi-flexible, or a flexible body. One of the major emerging trends in the later approach is to include the experimental information in the docking calculation. To this aim, this subproject deals with rigid body refinement on a complex formed by two proteins under ESR measured distance restraints. The refinement is performed by using a software package known as Crystallography and NMR System (CNS). This package has been traditionally used for macromolecular structure determination using crystallographic or NMR data as the input. It was necessary to make modifications in the program files in order to perform rigid body refinement on the basis of ESR distances. These distances were incorporated in a way similar to the NOE distances, which are in the form of a table and each distance (d) is allocated a negative error (dminus) and a positive error (dplus), thereby accounting for the flexibility of the spin label. The energy of the whole system is then minimized using the method of conjugate gradients. The refinement apart from depending on the accuracy of ESR measured distances, relies on the allocation of errors associated with each distance. At present, the program has been tested on a complex formed between chemotaxis proteins CheW and the P5 domain of delta 289 between which several ESR distances have been measured. Rigid body refinement gives a conformation of the two proteins that is close to the crystal structure of the complex. Significant is the fact that only 12 ESR distances between the two proteins were required to get a good approximation of their complex structure without any other supporting data. However, the program needs to be tested on more such systems, and a systematic way of assigning the errors associated with each ESR distance needs to be devised.