Butler J, Koppenol W H, Margoliash E
J Biol Chem. 1982 Sep 25;257(18):10747-50.
The temperature and pH dependence of the reaction of the superoxide radical anion with ferricytochrome c have been measured using the pulse-radiolysis technique. The temperature dependence of the reaction at low ionic strength yields an activation energy of 31 +/- 5 kJ/mol as compared to 14 +/- 3 kJ/mol for the reaction of CO2.(-) under the same conditions. The pH dependence fits the single pK'a of ferricytochrome c of 9.1. The bimolecular rate constant for the reaction of the superoxide anion with ferricytochrome c at pH 7.8, 21 +/- 2 degrees C, in the presence of 50 mM phosphate and 0.1 mM EDTA is (2.6 +/- 0.1) X 10(5) M-1 s-1. Using this value, 1 unit of superoxide dismutase activity (McCord, J. M., and Fridovich, I. (1969) J. Biol. Chem. 244, 6049-6055) is calculated to be 3.6 +/- 0.3 pmol of enzyme if the assay is performed in a total volume of 3.0 ml. Copper ions reduce the yield of the reaction of ferricytochrome c with CO2.(-). The reactivities of native and singly modified 4-carboxy-2,4-dinitrophenyllysine cytochromes c towards the superoxide anion radical are in the order native greater than 4-carboxy-2,4-dinitrophenyllysine 60 greater than lysine 13 greater than lysine 87 greater than lysine 27 greater than lysine 86 greater than lysine 72, indicating that electron transfer takes place at or close to the solvent accessible heme edge. The mechanism of the reaction is discussed in terms of the approach of superoxide anion radicals to the heme edge and the available molecular orbitals of both heme and free radicals.
利用脉冲辐解技术测定了超氧阴离子自由基与高铁细胞色素c反应的温度和pH依赖性。在低离子强度下，该反应的温度依赖性产生的活化能为31±5 kJ/mol，而在相同条件下，CO2(-)反应的活化能为14±3 kJ/mol。pH依赖性符合高铁细胞色素c的单一pKa值9.1。在pH 7.8、21±2℃、存在50 mM磷酸盐和0.1 mM EDTA的条件下，超氧阴离子与高铁细胞色素c反应的双分子速率常数为(2.6±0.1)×10(5) M-1 s-1。使用该值，如果在3.0 ml的总体积中进行测定，1个超氧化物歧化酶活性单位（麦考德，J. M.，和弗里多维奇，I.（1969年）《生物化学杂志》244，6049 - 6055）计算为3.6±0.3 pmol酶。铜离子降低了高铁细胞色素c与CO2(-)反应的产率。天然的和单修饰的4 - 羧基 - 2,4 - 二硝基苯赖氨酸细胞色素c对超氧阴离子自由基的反应活性顺序为天然的＞4 - 羧基 - 2,4 - 二硝基苯赖氨酸60＞赖氨酸13＞赖氨酸87＞赖氨酸27＞赖氨酸86＞赖氨酸72，表明电子转移发生在溶剂可及的血红素边缘或其附近。根据超氧阴离子自由基接近血红素边缘的方式以及血红素和自由基的可用分子轨道讨论了反应机理。
