The electronic state of heme in cytochrome oxidase II. Oxidation-reduction potential interactions and heme iron spin state behavior observed in reductive titrations.
The electronic state of heme in cytochrome oxidase II. Oxidation-reduction potential interactions and heme iron spin state behavior observed in reductive titrations.
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细胞色素氧化酶 II 中血红素的电子状态。
DOI：
10.1016/s0021-9258(17)38088-2
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发表时间：
1978
期刊：
影响因子：
--
通讯作者：
G. Palmer
中科院分区：
文献类型：
--
作者：
G. Babcock;L. Vickery;G. Palmer
磁圆二色性（MCD），电子顺磁共振（EPR），和光学吸收光谱已被用来监测氧化和还原血红素和铜的浓度在化学计量还原滴定纯化牛心细胞色素氧化酶。对MCD数据进行去卷积，以获得滴定期间还原细胞色素a和a3的浓度; EPR光谱分析提供了EPR可检测物质浓度的补充数据。对于不存在外源配体的天然酶，细胞色素a和a3在滴定中的所有点处被减少至大致相同的程度。另一方面，EPR可检测的铜的还原最初滞后于两种细胞色素的还原，但在滴定的最后阶段，在细胞色素a或a3之前完全还原。这些非能斯特滴定结果被解释为表明，在细胞色素氧化酶的血红素-血红素相互作用的主要模式涉及的两个细胞色素的氧化还原电位的变化，使氧化态的变化之一的血红素降低约135 mV的第二血红素的氧化还原电位。在这些滴定中，检测到最大占0.25自旋/氧化酶的高自旋物质。有证据表明，这些信号中的至少一些可以归因于细胞色素a3+，其在滴定过程中经历了低自旋到高自旋状态的转变。在一氧化碳的存在下，细胞色素a和a3的氧化还原性质明显改变。A32+。CO络合物在细胞色素a3+或EPR可检测的铜还原之前完全形成。g = 3的EPR信号归因于细胞色素a3+的细胞色素a2+的MCD强度的增加而减少;没有显着的高自旋强度观察到在任何中间阶段的还原。我们解释这些能斯特滴定结果表明，在配体的存在下，细胞色素a相对于细胞色素a3的氧化还原电位是由稳定的细胞色素a3配体复合物的氧化还原状态决定的，如果配体结合发生在还原的细胞色素a3上，则细胞色素a的电位较低;如果氧化的细胞色素A3通过配体结合而稳定，则具有更高电位的细胞色素A的结合。
Magnetic circular dichroism (MCD), electron paramagnetic resonance (EPR), and optical absorption spectroscopies have been used to monitor the concentrations of oxidized and reduced heme and copper during stoichiometric reductive titrations of purified beef heart cytochrome oxidase. The MCD data are deconvoluted to obtain the concentrations of reduced cytochromes a and a3 during the titrations; analysis of the EPR spectra provides complementary data on the concentrations of the EPR-detectable species. For the native enzyme in the absence of exogenous ligands, cytochromes a and a3 are reduced to approximately the same extent at all points in the titration. The reduction of the EPR-detectable copper, on the other hand, initially lags the reduction of the two cytochromes but in the final stages of the titration is completely reduced prior to either cytochrome a or a3. These non-Nernstian titration results are interpreted to indicate that the primary mode of heme-heme interaction in cytochrome oxidase involves shifts in oxidation-reduction potential for each of the two cytochromes such that a change in oxidation state for one of the hemes lowers the oxidation-reduction potential of the second heme by approximately 135 mV. In these titrations high spin species are detected which account for 0.25 spin/oxidase maximally. Evidence is presented to indicate that at least some of these signals can be attributed to cytochrome a3+ which has undergone a low-spin to high-spin state transition in the course of the titration. In the presence of carbon monoxide the oxidation-reduction properties of cytochromes a and a3 are markedly altered. The a32+. CO complex is fully formed prior to reduction of either cytochrome a3+ or the EPR-detectable copper. The g = 3 EPR signal attributed to cytochrome a3+ decreases as the MCD intensity of cytochrome a2+ increases; no significant high-spin intensity is observed at any intermediate stage of reduction. We interpret these Nernstian titration results to indicate that in the presence of ligands the oxidation-reduction potential of cytochrome a relative to cytochrome a3 is determined by the oxidation-reduction state of the stabilized cytochrome a3 ligand complex; if ligand binding occurs to reduced cytochrome a3 then cytochrome a titrates with a lower potential; cytochrome a titrates with a higher potential if oxidized cytochrome a3 is stabilized by ligand binding.