Proton-dependent electron transfer from CuA to heme a and altered EPR spectra in mutants close to heme a of cytochrome oxidase.
Proton-dependent electron transfer from CuA to heme a and altered EPR spectra in mutants close to heme a of cytochrome oxidase.
复制标题
质子依赖性电子从 CuA 转移到血红素 a，并改变了细胞色素氧化酶血红素 a 附近突变体的 EPR 谱。
DOI：
10.1021/bi801156s
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发表时间：
2008
期刊：
影响因子：
2.9
通讯作者：
Ferguson-Miller,Shelagh
中科院分区：
文献类型：
--
作者：
Mills,DeniseA;Xu,Shujuan;Geren,Lois;Hiser,Carrie;Qin,Ling;Sharpe,MartynA;McCracken,John;Durham,Bill;Millett,Francis;Ferguson-Miller,Shelagh
真核细胞色素氧化酶（CcO）与杜鹃花和拟杜鹃花的同源原核形式在血红素的EPR谱中存在差异。注意到血红素（H102）的组氨酸配体与Rhodobacter（S44）和ParaccocusCc 0中的丝氨酸氢键结合，而与牛酶中的甘氨酸相反。S44突变为甘氨酸使hemeaEPR信号从gz= 2.82转变为2.86，更接近牛hemeaat 3.03，而不改变其他性质。然而，突变为天冬氨酸，导致相反的移位和分裂hemeaEPR信号的gz= 2.72/2.78，伴随着较低的活性和急剧抑制的内在电子转移从CuA血红素。这种内在速率是双相的;慢的比例是pH依赖性的，这是两个EPR信号成分的相对强度。在pH 8时，2.72处的hemeaEPR信号最强，电子转移速率（CuA到hemea）为10−130 s−1，而野生型为90000 s−1。在pH 5.5时，2.78处的信号增强，并且观察到双相速率，50%快（野生型）和50%慢（90 s-1）。这些数据支持的预测，组氨酸配体的血红素的氢键合作伙伴是一个决定因素的EPR光谱之间的差异牛和细菌CcO。我们进一步证明，hemearedox电位可以显着改变附近的羧基，其质子化导致质子耦合的电子转移过程。
Eukaryotic cytochromecoxidase (CcO) and homologous prokaryotic forms ofRhodobacterandParaccocusdiffer in the EPR spectrum of hemea. It was noted that a histidine ligand of hemea(H102) is hydrogen bonded to serine inRhodobacter(S44) andParaccocusCcOs, in contrast to glycine in the bovine enzyme. Mutation of S44 to glycine shifts the hemeaEPR signal from gz= 2.82 to 2.86, closer to bovine hemeaat 3.03, without modifying other properties. Mutation to aspartate, however, results in an oppositely shifted and split hemeaEPR signal of gz= 2.72/2.78, accompanied by lower activity and drastically inhibited intrinsic electron transfer from CuAto hemea. This intrinsic rate is biphasic; the proportion that is slow is pH dependent, as is the relative intensity of the two EPR signal components. At pH 8, the hemeaEPR signal at 2.72 is most intense, and the electron transfer rate (CuAto hemea) is 10−130 s−1, compared to wild-type at 90000 s−1. At pH 5.5, the signal at 2.78 is intensified, and a biphasic rate is observed, 50% fast (∼wild type) and 50% slow (90 s−1). The data support the prediction that the hydrogen-bonding partner of the histidine ligand of hemeais one determinant of the EPR spectral difference between bovine and bacterial CcO. We further demonstrate that the hemearedox potential can be dramatically altered by a nearby carboxyl, whose protonation leads to a proton-coupled electron transfer process.