THERMODYNAMIC AND KINETIC STUDIES OF CYTOCHROME C OXIDASE

细胞色素C氧化酶的热力学和动力学研究

• 批准号: 3843254
• 负责人: R W HENDLER
• 金额: --
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3843254
• 关键词: Raman spectrometry; atomic absorption spectrometry; bacteriorhodopsins; bioenergetics; biomedical equipment development; cellular respiration; cytochrome oxidase; enzyme activity; isozymes; oxidation reduction reaction; photochemistry; potentiometry; thermodynamics

Resonance Raman spectroscopy has been combined with the electric voltage control techniques developed in this laboratory to add a new dimension to the potentiometric characterization of cytochrome aa3. This new approach confirms the existence (previously established in this laboratory) of a low potential species of cytochrome a3 (Em=200 mV), and of two forms of cytochrome a (Em's of 260 and 340 mV). In addition, a new form of cytochrome a3 (Em=approximately 260 mV) has been found. The technique also showed that the absorbance at 830 nm is not a unique monitor for CuA, as has been assumed. The new ultrarapid spectrometer, under development in the laboratory, has been completed. Preliminary studies on the reduction of cytochrome aa3 by cytochrome c, indicate a more complicated series of electron transfer events than are currently believed. Two distinct kinetic forms of the enzyme can be observed depending on the presence or absence of a small amount of O2. Multichannel studies on the laser- activated photocycle of bacteriorhodopsin have been initiated. The kinetic spectra obtained have been deconvoluted by SVD and it has been confirmed that the level of laser activation does influence the kinetics of the ensuing photocycle, changing the relative weights of key intermediates. A new optical bench setup has been constructed that vastly improves the sensitivity of the system, allowing for a very low level of monitoring light, and having the ability to detect absorbance changes in the submilli-O.D. range.

共振拉曼光谱已与电压相结合 该实验室开发的控制技术为 细胞色素 aa3 的电位表征。这种新方法 确认存在（之前在本实验室建立的） 细胞色素 a3 的低电位种类 (Em=200 mV)，以及两种形式 细胞色素 a（Em 为 260 和 340 mV）。此外，还有一种新形式 已发现细胞色素a3（Em=约260 mV）。该技术还 表明 830 nm 处的吸光度并不是 CuA 的唯一监测器，因为 已被假设。新型超快速光谱仪，正在开发中 实验室，已建成。减量的初步研究 细胞色素 aa3 与细胞色素 c 的结合，表明一系列更复杂的 电子转移事件比目前认为的要多。 两种截然不同的 可以观察到酶的动力学形式，具体取决于酶的存在或 缺乏少量O2。 激光多通道研究 细菌视紫红质的激活光循环已启动。动力学 获得的光谱已通过 SVD 解卷积并得到证实 激光激活水平确实影响动力学 随后发生光循环，改变关键中间体的相对重量。一个 新的光具座装置已经建成，极大地改善了 系统的灵敏度，允许非常低水平的监控 光，并具有检测吸光度变化的能力 亚毫外径范围。