Metalloenzyme Mechanisms

超出提供的空间。通过这项研究应用的广泛的、长期的目标，我们试图了解生物大分子在形成大分子组件时识别其伙伴的机制。根据以下目标，建议对生物系统中的识别事件的主要类别进行系统研究。(1)蛋白质-蛋白质识别。在这里，我们试图定义特定表面相互作用的作用，即通过表面互补提供的静电、氢键和疏水自由能，它们定义了在真核细胞色素P450依赖的加氧酶中协调电子转移事件的金属蛋白之间形成络合物的特异性和亲和力。(2)膜成分识别。生物膜具有由磷脂、脂肪酸、胆固醇和其他关键细胞成分组成的复杂结构，不仅仅是嵌入真核细胞P450蛋白成分的简单“洗涤剂”。双层的组成和物理性质对于作为目标(1)的主题的识别事件是重要的。为了了解膜结构对分子识别的贡献，多种技术应运而生。(3)蛋白质-小分子识别。在这个目标中，我们试图确定相同的静电学、氢键和底物与酶活性部位的手手套匹配的基本力如何能够在细胞色素P450催化中引起观察到的区域和立体特异性的高度控制。(4)蛋白质-核酸识别。同样，同样的基本力量控制着识别过程。在这一目标中，我们在以前成功定义了溶剂水在蛋白质和核酸成分之间的氢键识别中的作用方面取得了进展。我们通过研究有趣的“间接读出”大分子组件来扩展系统的研究范围，并使用新的流体静力学和渗透压方法，与结晶学和分子动力学模拟相结合。在这次竞争性更新中，我们建议的研究计划协同使用分子生物学中广泛的跨学科工具和技术以及先进的生物物理方法，这些方法已被证明是了解金属酶机制的理想方法。PERFORMANCESITE(========================================Section End===========================================

EXCEED THE SPACE PROVIDED. Through the broad, long term objectives of this research application, we seek to understand the mechanisms by which biological macromolecules recognize their partners in the formation of macromolecular assemblies. Systematic investigations are proposed that focus on the major classes of recognition events in biological systems according to the following Aims. (1) Protein - Protein Recognition. Here we seek to define the role of specific surface interactions; electrostatic, hydrogen bonding, and hydrophobic free energies provided through surface complimentary, which define the specificity and affinity in the formation of complexes between the metalloproteins coordinating electron transfer events in the eukaryotic cytochrome P450 dependent oxygenases. (2) Membrane component recognition. Biological membranes, with their complex makeup of phospholipids, fatty acids, cholesterol and other critical cellular components, is more than a simple "detergent" for embedded eukaryotic P450 protein components. The composition and physical properties of the bilayer are important for the very recognition events that are the subject of Aim (1). Multiple techniques are brought to bear toward the goal of understanding the contributions of membrane architectures to molecular recognition. (3) Protein - Small Molecule Recognition. In this Aim we seek to ascertain how the same fundamental forces of electrostatics, hydrogen bonding and the hand - glove fit of a substrate into the active site of an enzyme can give rise to the observed high degree control of regio- and stereo- specificity in cytochrome P450 catalysis. (4) Protein - Nucleic Acid Recognition. Again the same fundamental forces control recognition processes. In this Aim we build on previous success in defining roles of solvent water in mediating hydrogen bond recognition between protein and nucleic acid components. We expand the repertoire of systems investigated by examining the interesting class of "indirect readout" macromolecular assemblies and use novel hydrostatic and osmotic pressure methods, in concert with crystallography and molecular dynamics simulations. Our proposed research program in this competitive renewal make concerted use of broad interdisciplinary tools and techniques in molecular biology and advanced biophysical methods which have proven to be ideal for understanding the fundamental mechanisms of metalloenzyme mechanisms. PERFORMANCESITE( ========================================Section End===========================================