Molecular mechanism of redox driven proton pumps

电子和质子的耦合转移被认为是一次能源的普遍原理 保持呼吸。末端呼吸氧化酶是膜结合的电子- 催化氧还原为水的转移络合物，该反应与 随着跨膜质子梯度的产生，跨膜质子梯度是细胞的主要来源， 自由能源氧化酶利用两个基本原理来产生质子梯度。第一个是 经典的米切尔环机制，其中用于水形成的电子和质子是 从膜的相对侧吸收。第二个是质子泵， 通过氧化还原反应提供的自由能驱动质子穿过膜的转移。 然而，质子泵的质子泵送机制还没有在任何质子泵中阐明 它仍然是分子生物学的关键问题之一， 生物能量学拟议的调查将有助于了解质子 通过鉴定电子-质子耦合位点和 实验研究和建立的机械原理的泵和 血红素-铜氧化酶中的质子门控。光学光谱学、电子 顺磁共振，快速冷冻淬灭，闪光光解，停流，和几个 将利用生物化学方法来实现这些目标。将讨论具体目标 使用纯化的牛心氧化酶，将酶掺入磷脂囊泡中， 亚线粒体颗粒

Coupled electron and proton transfer is recognized as the universal principle of primary energy conservation in respiration. Terminal respiratory oxidases are membrane-bound electron- transfer complexes that catalyze the reduction of oxygen to water and this reaction is associated with the generation of a transmembrane proton gradient that is the primary source of cellular free energy. Oxidases utilize two basic principles to produce the proton gradient. The first is a classical Mitchell's loop mechanism in which electrons and protons for the water formation are taken up from the opposite sides of membrane. The second one is proton pumping in which proton transfer across the membrane is driven by the free energy provided by a redox reaction. However, the mechanism of proton pumping has not been elucidated in any proton pump driven by reduction-oxidation reactions and it remains one of the key problems of molecular bioenergetics. Proposed investigations will contribute to an understanding of the proton translocation mechanism by identification of the electron-proton coupling sites and experimental examination and establishment of the mechanistic principles of pumping and proton gating in heme-copper oxidases. The application of optical spectroscopy, electron paramagnetic resonance, rapid freeze-quenching, flash photolysis, stopped-flow, and several biochemical methods will be utilized to achieve these goals. Specific aims will be addressed using the purified bovine heart oxidase, enzyme incorporated into phospholipid vesicles and in submitochondrial particles.