Sato K, Hasumi H, Tsukidate A, Sakurada J, Nakamura S, Hosoya T
Faculty of Pharmaceutical Sciences, Chiba University, Japan.
Biochim Biophys Acta. 1995 Nov 15;1253(1):94-102. doi: 10.1016/0167-4838(95)00144-j.
The effects of methanol, acetone, and ethylene glycol (up to 50% v/v) on elementary steps in the reactions of horseradish peroxidase (HRP) and lactoperoxidase (LPO) were studied by means of the stopped-flow method and the difference spectrum. The rate constant (k3,app) of the oxidation reaction of p-cresol with HRP compound II was remarkably reduced in the presence of organic solvents (to 2.3%, 1.8% and 9.4% of the original value in the presence of 50% (v/v) of methanol, acetone and ethylene glycol, respectively), then to a lesser degree were decreased the rate of the oxidation reaction with LPO compound II, and the rate of the oxidation reaction with HRP compound I. These reductions in the reaction rates were not due to competitive inhibition of the solvents, but considered to be related to the degree of exposure of the electron transfer route to the medium. While the rate constant of compound I formation (k1,app) was moderately affected by organic solvents in the case of HRP, the reaction rate with LPO was scarcely affected by organic solvents, being in harmony with the compact heme crevice which probably hampers penetration of solvent molecules. The rate constant (k2,i,app) of the oxidation reaction of an iodide ion by HRP compound I was also hardly affected by the solvents. On the basis of these facts, the mechanism of electron transfer from donors to compound I and compound II is discussed.
采用停流法和差示光谱法研究了甲醇、丙酮和乙二醇（体积分数高达50%）对辣根过氧化物酶（HRP）和乳过氧化物酶（LPO）反应基本步骤的影响。在有机溶剂存在下，对甲酚与HRP化合物II的氧化反应速率常数（k3,app）显著降低（在50%（v/v）甲醇、丙酮和乙二醇存在下，分别降至原始值的2.3%、1.8%和9.4%），其次是与LPO化合物II的氧化反应速率，以及与HRP化合物I的氧化反应速率降低程度较小。反应速率的这些降低并非由于溶剂的竞争性抑制，而是被认为与电子转移途径暴露于介质的程度有关。虽然在HRP的情况下，化合物I形成的速率常数（k1,app）受到有机溶剂的适度影响，但与LPO的反应速率几乎不受有机溶剂的影响，这与可能阻碍溶剂分子渗透的紧密血红素裂隙相一致。HRP化合物I氧化碘离子的反应速率常数（k2,i,app）也几乎不受溶剂的影响。基于这些事实，讨论了从供体到化合物I和化合物II的电子转移机制。
