Rapid Scanning Stopped Flow Spectrophotometer for Enzymology

将对参与芳烃代谢、烷基胺降解和呼吸作用的酶进行机制和动力学研究。这些酶包括黄色素对甲酚甲基羟化酶（恶臭假单胞菌）、醌蛋白甲胺脱氢酶（甲基营养菌）、含铜醌蛋白胺氧化酶（人和球形节杆菌）、铁硫黄素蛋白琥珀酸脱氢酶（哺乳动物和细菌）和大肠杆菌的延胡索酸还原酶。已经定义了有机和金属离子辅因子的数量、类型和氧化还原电位。 这些酶中的大多数已被克隆，以允许通过遗传操作改变其特性。许多研究蛋白质的可用X射线结构将指导一些研究。建议使用所要求的快速扫描停流分光光度计来定义底物/配体与酶结合时发生的事件，以及集成功能中单个辅因子的特性。它还建议，该仪器可用作快速记录，双波长分光光度计基本醌醇氧化还原测定。

Mechanistic and kinetic studies of enzymes involved in the metabolism of aromatic hydrocarbons, degradation of alkylamines, and respiration will be undertaken. The enzymes include the flavocytochrome p-cresol methylhydroxylase (Pseudomonas putida), the quinoprotein methylamine dehydrogenase (methylotrophic bacteria), copper-containing quinoprotein amine oxidases (human and Arthrobacter globoformis), and iron-sulfur flavoproteins succinate dehydrogenase (mammalian and bacteria) and fumarate reductase from Escherichia coli. The numbers, types, and redox potentials of the organic and metal ion cofactors are already defined. The majority of these enzymes have been cloned to allow for alteration of their properties by genetic manipulation. The available x-ray structures of many of the proteins studied will guide some of the research. It is proposed that the requested rapid-scanning stopped-flow spectrophotometer be used to define events occurring on substrate/ligand binding to enzymes, and the properties of the individual cofactors in integrated functions. It is also proposed that the instrument be used as a rapid-recording, dual-wavelength spectrophotometer for essential quinol oxidation reduction assays.