Fundamentals of Ligand-Protein Interactions

配基-蛋白质复合体的小分子配基的构象变化是许多药物作用的中心方面，也是药物设计计算方法中的一个关键挑战。在该领域最早的出版物之一中，我们对蛋白质数据库(PDB)和剑桥结构数据库(CSD)中出现的一小部分配体表明，柔性化合物通常不会以其全局真空能量构象与蛋白质结合，甚至在任何局部真空能量构象中也不会。虽然这项研究使用了当时可用的最大数据集和最佳方法，但自那以来，实验数据库中的结构数量以及可用的软件和硬件资源都呈指数级增长。因此，我们正在重新讨论这个重要的话题，对数量级的分析，更多的结构，以及在计算量子化学理论的高水平上执行的计算。在这个项目到目前为止取得的其他里程碑中，我们提取了最近在PDB的LigandEXPO中提供的所有小分子配体的出现。截至2008年5月，这是一组超过350,000组不同的3D坐标(当前组大小：190万)。我们添加了来自几个不同来源的大量注释。在过滤器链中使用这些注释，我们已经生成了高质量和高可靠性的配体结构的“高质量”子集，根据应用的严格程度，从大约1000次到大约5000次不等。我们对这些高质量配体的构象能进行了高水平的量子化学计算。在第一轮中，真空能量计算部分在我们自己的Linux集群上运行，部分在美国国立卫生研究院CIT的Biowulf集群上运行。在这个计算量巨大的项目中，同时使用了多达1000个CPU，单个作业需要几个小时到几个星期的CPU时间。我们从大约360次成功完成的运行中获得了结果。这些结果清楚地表明，这些量子化学计算充分证实了高构象能的可能性。为了探索水环境对配体构象能的可能影响-毕竟，真空并不是药物分子通常操作的地方-进行了第二轮量子化学计算，使用了Gauss03中的SCI-PCM溶剂模型。与真空计算相比，这些运行对计算机资源的要求更高。为了分析能量不确定性作为位置不确定性的函数，而位置不确定性又是结晶学分辨率的函数，我们在分子力学力场水平上以晶体结构构象为中心进行了构象采样，得到了依赖于分辨率的扭转分布。更强大的实验仪器的出现，如自由电子激光，为回答这些问题开辟了新的可能性。与这一主题相关的是最近开始的一项关于有机小分子互变异构的研究，这是化学信息学和数据库中的一个重要问题(项目3)，高效药物设计(项目2和3)，以及目前更好地了解蛋白质-配体相互作用和帮助这一探索的晶体结构的项目。这项工作由劳拉·瓜施-帕米斯博士完成。互变异构的工作仍在继续，综合实验、量子化学和化学信息学分析的有趣结果已在已接受或正在审查的论文中描述，和/或为进一步分析而研究。这一领域的工作为涉及醛糖还原酶和其他癌症和HPV相关蛋白的晶体结构的令人兴奋的新研究奠定了基础。该项目发表了一些评论该领域的现状和未来可能性的论文，包括最近发表的超高分辨率结晶学的作用。随着CADD小组工作中越来越重视对接，这一专题获得了新的重要性。

The conformational changes of both partners of a ligand-protein complex, the small-molecule ligand in its the protein binding site (in many cases the catalytically active site of an enzyme) are a central aspect many drug actions, as well as a crucial challenge in computational approaches to drug design. In one of the earliest publications in the this field, we showed for a small set of ligands occurring both in the Protein Data Bank (PDB) and the Cambridge Structural Database (CSD) that flexible compounds are not usually bound to a protein in their global vacuum energy conformation, and oftentimes not even in any local vacuum energy conformation. While this study used the largest set of data and best methodology available at that time, both the number of structures in either experimental database and the software and hardware resource available have since grown exponentially. We are thus revisiting this important topic with an analysis of orders of magnitudes more structures, and computations performed at a high level of computational quantum-chemical theory. Among other milestones achieved so far in this project, we have extracted all occurrences of small-molecule ligands recently made available in PDB's LigandExpo. As of May 2008, this was a set of over 350,000 distinct sets of 3D coordinates (current set size: 1.9 million). We have added extensive annotation coming from several different sources. Using these annotations in a chain of filters, we have generated "high-quality" subsets of ligand structures of high quality and reliability numbering from just about one thousand to about 5,000 occurrences depending on the stringency applied. We have conducted high-level quantum-chemical calculations of conformational energies for these high-quality ligand sets. In the first round, vacuum energy calculations were run partly on our own Linux cluster, partly on the Biowulf cluster of the CIT, NIH. Up to a thousand CPUs were used simultaneously in this computationally massive project, with individual jobs taking from a few hours to several weeks of CPU-time. We obtained results from about 360 runs that completed successfully. These results clearly showed that the possibility for high conformational energies are fully confirmed by these quantum-chemical calculations. To explore the possible influence of aqueous environment on ligand conformational energies - after all, vacuum is not really where drug molecules typically operate - a second round of quantum chemical calculations was conducted, employing the SCI-PCM solvent model in Gaussian 03. These runs were even more demanding in terms of computer resources than the vacuum calculations. To analyze the energetic uncertainty as a function of the positional uncertainty, which in turn is a function of the crystallographic resolution, we conducted sampling of conformations with a resolution-dependent torsion distribution centered around the crystal structure conformation at the molecular mechanics force field level. The advent of ever more-powerful experimental instrumentation such as free electron lasers opens up new possibilities in answering these questions. Related to this topic is a study recently begun on tautomerism of small organic molecules, which is an important question both in chemoinformatics and databases (Project 3), efficient drug design (Projects 2 and 3), and the present project of better understanding protein-ligand interactions and the crystal structures aiding in this quest. This work has been performed by Dr. Laura Guasch-Pamies. The tautomerism work is continuing with interesting results of combined experimental, quantum-chemical and chemoinformatics analysis, which have been described in papers either accepted or under review, and/or studied for further analyses. The work in this field has laid the groundwork for exciting new research involving crystal structures of aldose reductase and other cancer- and HPV-related proteins. A number of papers reviewing the current status and the future possibilities of the field have been published in this project, including most recently the role of ultra-high resolution crystallography. This topic has gained new importance with the increased emphasis of docking in the CADD Group's work.