Biophysical Studies of Metalloenzymes

该项目包括两个方面的工作：金属酶的电子-核双共振(Endor)研究和蛋白质复合体内远程电子转移的研究。Endor实验提供了那些与顺磁中心的电子自旋相互作用的原子核的核磁共振谱，通过适当的同位素标记，它可以表征金属酶活性中心的每个原子。蛋白质-蛋白质复合体中的长程电子转移可以通过用锌或镁原卟啉取代其中一个配对的血红素来研究。电子顺磁共振(EPR)技术是研究金属蛋白结构和功能的重要手段。在很大程度上，EPR提供的关于金属中心的组成、结构和成键的信息包含在金属核、内源配体和结合底物的电子-核超精细耦合中。遗憾的是，对于大多数矿物生物分子，超精细裂解在普通EPR中是不能分辨的，并且丢失了信息。恩多光谱学恢复了这一信息。电子转移反应是生物和化学的核心，但直到最近才发展出研究远程生物电子转移的技术，通过使用修饰的蛋白质在固定距离保持电子供体/受体氧化还原对来研究远程生物电子转移，而不会使二级过程复杂化。

This project comprises two efforts: electron-nuclear double resonance (endor) investigations of metalloenzymes and studies of long-range electron transfer within protein complexes. An endor experiment provides an nmr spectrum of those nuclei that interact with the electron spin of a paramagnetic center, and with appropriate isotopic labelling it can characterize every atom of a metalloenzyme active site. Long-range electron transfer within a protein-protein complex can be studied by replacing the heme of one partner with a zinc or magnesium protoporphyrin. Electron paramagnetic resonance (epr) techniques are of major importance in the study of metalloprotein structure and function. In large part, the information epr provides about the composition, structure and bonding of a metal center is contained in the electron-nuclear hyperfine couplings of the metal nuclei, endogenous ligands and bound substrates. Unfortunately, for most metallobiomolecules the hyperfine splittings are not resolvable in ordinary epr and the information is lost. Endor spectroscopy recovers this information. Electron transfer reactions are central to biology and chemistry, but only recently have techniques been developed to study long- range biological electron transfer without the complication of second order processes, through use of modified proteins which hold an electron donor/acceptor redox pair at fixed distance.