Electrochemical evidence for multiple peroxidatic heme states of the diheme cytochrome c peroxidase of Pseudomonas aeruginosa.
Electrochemical evidence for multiple peroxidatic heme states of the diheme cytochrome c peroxidase of Pseudomonas aeruginosa.
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DOI：
10.1021/bi801699m
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发表时间：
2009-01-13
期刊：
影响因子：
2.9
通讯作者：
Elliott, Sean J.
中科院分区：
文献类型：
--
作者：
Becker, Clinton F.;Watmough, Nicholas J.;Elliott, Sean J.
采用蛋白膜伏安法研究了铜绿假单胞菌细胞色素c过氧化物酶及其催化机理。双血红素细菌酶的单分子膜被固定在热解石墨边缘和烷醇修饰的Au电极上。与低电位血红素相关的氧化还原对可以在两个电极表面上检测到，还原电位为−234 mV vs SHE。的中点电位显示在酸性pH值，指示质子耦合电子转移的一个独特的pH依赖性。LP血红素的非翻转信号在加入底物后可转变为S形波。使用转换信号的中点电位计算米氏动力学，Km = 25 μM。加入氰化物（Ki = 50 μM）可抑制催化作用。这些动力学参数与先前报道的基于溶液的研究结果吻合良好，表明酶的活性不受电极表面固定化的影响。催化波的还原电位清楚地表明，在电催化过程中的限速物种不同于以前报道的过氧化物酶，这表明PFV可用于在未来区分细菌细胞色素c过氧化物酶的还原活化的要求。
The enzyme cytochrome c peroxidase from Pseudomonas aeruginosa and its catalytic mechanism was investigated using protein film voltammetry. Monolayers of the diheme bacterial enzyme were immobilized on both pyrolytic graphite edge and alkanethiol modified Au electrodes. The redox couple associated with the low potential heme could be detected on both electrode surfaces at a reduction potential of −234 mV vs SHE. The midpoint potential displays a distinct pH dependence at acidic pH values, indicative of proton-coupled electron transfer. The non-turnover signal of the LP heme can be transformed into sigmoidal waves upon the addition of substrate. The midpoint potential of the turnover signals were used to calculate Michaelis-Menten kinetics with a Km = 25 μM. Catalysis was inhibited with addition of cyanide (Ki = 50 μM). These kinetic parameters are in good agreement with previously reported solution-based studies, indicated that the activity of the enzyme is unaffected by the immobilization on the electrode surface. The reduction potential of the catalytic wave clearly shows that rate-limiting species during electrocatalysis differs from those previously reported for peroxidases, indicating that PFV may be used in the future to distinguish the requirement for reductive activation in bacterial cytochrome c peroxidases.
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影响因子：
3
作者：
de Sousa, P. M. Paes;Pauleta, S. R.;Moura, J. J. G.
通讯作者：
Moura, J. J. G.
影响因子：
3.9
作者：
Hsiao, Hsi-Chen;Boycheva, Svetlana;Brittain, Thomas
通讯作者：
Brittain, Thomas
DOI：
10.3891/acta.chem.scand.29b-0719
发表时间：
1975-01-01
期刊：
ACTA CHEMICA SCANDINAVICA SERIES B-ORGANIC CHEMISTRY AND BIOCHEMISTRY
影响因子：
--
作者：
RONNBERG, M;ELLFOLK, N
通讯作者：
ELLFOLK, N
影响因子：
2.9
作者：
Echalier, Aude;Brittain, Thomas;Watmough, Nicholas J.
通讯作者：
Watmough, Nicholas J.
DOI：
10.1016/0167-4838(83)90413-2
发表时间：
1983-01-01
期刊：
BIOCHIMICA ET BIOPHYSICA ACTA
影响因子：
--
作者：
ELLFOLK, N;RONNBERG, M;VANNGARD, T
通讯作者：
VANNGARD, T