CAREER: Correlating Metalloenzyme Structure with Reactivity By Tunneling Electrons in Crystals

归根结底，生命的本质是电荷的受控运动。多肽链的大量构象和化学状态允许自然调节金属中心的反应性和蛋白质内部和蛋白质之间的直接电子流动。Crane博士的研究目标是开发和应用新的光化学方法来研究生物中氧化还原化学和远程电子转移(ET)的结构基础。光敏剂将被用来在单个蛋白质晶体中引发远程ET，其中结构可以通过X射线结晶学精确定义。设计的固定供体和受体取向的晶体系统允许通过突变和同位素替代来探测介入蛋白质和水结构对电子隧穿的影响。ET率将直接在金属修饰的天青素晶体和细胞色素c与细胞色素c过氧化物酶络合物的晶体中确定。蛋白质金属中心的氧化还原反应将在光诱导ET的作用下在晶体中进行，因此对催化至关重要的活化态的结构可以通过低温结晶学或时间分辨衍射技术来确定。将确定过氧化物酶、过氧化铁和氧化铁物种的结构，这些物种是生物氧活化的重要中间体，以及色氨酸和酪氨酸自由基，它们是各种高潜力生物氧化还原化学中的新兴参与者。原子结构将与电子结构相关联，用磁光谱和光谱学来探测，以了解金属辅因子和多肽链之间的反应活性的相互调节。这项研究的结果将提供一组详细定义的结构状态的长程ET率，迄今未观察到的金属蛋白催化物种的新结构，以及通过X射线衍射研究蛋白质动力学的新技术。与这项研究相关的教育活动旨在将结构、动力学、热力学和化学反应的基本概念与复杂的生物过程如能量转导、信号转导、环境敏感性和响应联系起来。努力分为两大类：1)指导高中生、本科生和研究生的研究；2)在康奈尔大学开发和教授新的本科生和研究生课程。还将促进K-12阶段的基于发现的学习。上述研究旨在为所有级别的学生提供合适的项目。目前正在教授一门新的研究生课程--“蛋白质的物理化学”，第二门课程“生物无机化学”将于2003年推出。其他课程活动包括为化学专业一年级学生开发一门化学结构和成键入门课程，以及一门跨部门的酶动力学短期课程。在所有活动中都将强调有趣的方法来解决问题。

Ultimately, the essence of life is the controlled movement of charge. The vast number of conformational and chemical states available to the polypeptide chain allows nature to tune the reactivity of metal centers and direct electron flow within and between proteins. The goal of Dr. Crane's research is to develop and apply new photochemical methods for studying the structural basis of redox chemistry and long-range electron transfer (ET) in biology. Photosensitizers will be used to initiate long-range ET in single protein crystals, where structure can be precisely defined by X-ray crystallography. Designed crystal systems that fix donor and acceptor orientations allow effects of intervening protein and water structure on electron tunneling to be probed by mutagenesis and isotopic substitution. ET rates will be determined directly in crystals of metal-modified azurins and crystals of complexes between cytochrome c and cytochrome c peroxidase. Redox reactions at protein metal centers will be driven in crystals by photoinduced ET so that structures of activated states important for catalysis can be determined by cryo-crystallography or time-resolved diffraction techniques. Structures will be determined for peroxidase peroxo-iron and oxo-iron species, important intermediates in biological oxygen activation, and for tryptophan and tyrosine radicals, emerging players in a wide variety of high-potential, biological redox chemistries. Atomic structure will be correlated with electronic structure probed with magnetic and optical spectroscopies to understand the reciprocal tuning of reactivity between metallocofactors and the polypeptide chain. The results of this research will provide sets of long-range ET rates for structural states defined in detail, new structures of hitherto unobserved metalloprotein catalytic species, and new techniques for studying protein dynamics by X-ray diffraction.Educational activities connected with this research aim to relate fundamental concepts in structure, kinetics, thermodynamics and chemical reactivity to complex biological processes such as energy transduction, signal transduction, environmental sensitivity and response. Effort falls into two general categories: 1) mentoring high-school, undergraduate, and graduate students in research; and 2) the development and teaching of new undergraduate and graduate courses at Cornell University. Discovery-based learning at the K-12 level will also be promoted. The research described above is designed to provide projects appropriate for students of all levels. A new graduate course, " the physical chemistry of proteins" is now being taught and a second "Bio-inorganic chemistry" will be presented in 2003. Other course activities include development of an introductory course for freshman chemistry majors in chemical structure and bonding, and an interdepartmental short course in enzyme kinetics. A playful approach to addressing problems will be emphasized in all activities.