Mann A J, Auer H E
J Biol Chem. 1980 Jan 25;255(2):454-8.
Purified beef heart cytochrome c oxidase is inactivated to the extent of 35 to 50% by the nonpolar mercurial reagents mercuric chloride and ethylmercuric chloride. The inactivation is complete within 5 min. In titrations of activity, the plateau level of inactivation is attained at added ethylmercuric chloride:heme a ratios of about 1:1. Up to 3 mercury atoms/heme a are bound to the oxidase, although only the first of these affects its enzymatic activity. Incubation of the ethylmercury-modified oxidase with sulfhydryl compounds reverses the inactivation, with 2,3-dimercaptopropanol being most effective of the reagents tested. Spectrophotometric and polarographic assays of enzymatic activity show that Km values for the native and the ethylmercury-modified enzymes are practically indistinguishable, and that the partial inactivation observed for the latter is reflected exclusively in a lower value of Vmax compared to that of the native enzyme. Based on these results, we propose that ethylmercuric chloride reacts with a single crucial--SH group per heme a, and that electron transfer processes in the modified product are partially inhibited.
纯化的牛心细胞色素c氧化酶会被非极性汞试剂氯化汞和乙基氯化汞灭活35%至50%。灭活在5分钟内完成。在活性滴定中，当加入的乙基氯化汞与血红素a的比例约为1:1时，达到灭活的平稳水平。每分子血红素a最多有3个汞原子与氧化酶结合，不过其中只有第一个汞原子会影响其酶活性。用巯基化合物孵育乙基汞修饰的氧化酶可使灭活逆转，在所测试的试剂中，2,3-二巯基丙醇最为有效。酶活性的分光光度法和极谱法测定表明，天然酶和乙基汞修饰酶的米氏常数实际上无法区分，后者观察到的部分失活仅表现为与天然酶相比最大反应速度值较低。基于这些结果，我们提出乙基氯化汞与每个血红素a的一个关键巯基反应，并且修饰产物中的电子转移过程受到部分抑制。
