Structurally Engineered Photoredox Enzymes

加州理工学院的Gray和Winkler博士将在化学部无机、生物无机和有机金属项目以及分子和生物化学科学部生物物理项目的支持下，研究修饰金属酶的光化学。这些工作人员将结合无机化学、物理化学和分子生物学的方法和系统，定制设计光驱动多电子氧化还原酶。它们会光引发单电子转移反应，导致在细胞色素c和肌红蛋白的变体中形成含氧铁（铁基）。这些蛋白质将通过使用半合成和定点诱变技术为铁基形成量身定制。钌结合的组氨酸残基将被引入蛋白质的表面位点，蛋白质血红素环境的改变将使蛋白质适应于支持活性位点的铁基物种。在光解过程中，钌和血红素之间的电子转移将导致铁基的形成。在许多由血红素蛋白催化的双氧反应中，一种重要的中间物质是含有血红素的铁原子与氧原子结合的中间物质。这些“铁基”物种在过去被证明是难以表征的，因为它们的高反应性。一种产生和检测它们的新颖而强大的方法将被开发出来，这首先依赖于使用分子生物学方法来设计蛋白质，以便于由短暂的激光闪光驱动的电子转移反应。然后利用它们的光学性质来检测铁基物质，并直接获得有关它们的反应性的信息。

Doctors Gray and Winkler of CalTech will investigate the photochemistry of modified metalloenzymes with support from the Inorganic, Bioinorganic, and Organometallic Program of the Chemistry Division and the Biophysics Program of the Molecular and Biochemical Sciences Division. These workers will combine methods and systems from inorganic chemistry, physical chemistry, and molecular biology to custom design light-driven multielectron redox enzymes. They will photoinitiate single-electron-transfer reactions that will result in formation of oxo-iron (ferryl) species in variants of cytochrome c and myoglobin. These proteins will be tailored for ferryl formation by use of semisynthetic and site-directed mutagenesis techniques. Histidine residues for ruthenium binding will be introduced at surface sites on the proteins, and alterations in the protein heme environments will adapt the protein to support ferryl species in the active sites. Upon photolysis, electron transfer between the ruthemium and heme moieties will lead to ferryl formation. %%% An important intermediate species in many reactions of dioxygen that are catalyzed by heme proteins is one that contains the iron atom of the heme bound to an oxygen atom. These "ferryl" species in the past have proven difficult to characterize because of their high reactivity. A novel and powerful method to generate and detect them will be developed that depends first on using methods of molecular biology to engineer the proteins so as to facilitate electron transfer reactions that can be driven by a short flash of laser light. The ferryl species will then be detected by use of their optical properties, and information on their reactivity will be obtained directly.