Hashimoto F, Hayashi H
Biochim Biophys Acta. 1987 Sep 4;921(1):142-50.
Catalase activity was inhibited by aminotriazole administration to rats in order to evaluate the influence of catalase on the peroxisomal fatty acyl-CoA beta-oxidation system. 2 h after the administration of aminotriazole, peroxisomes were prepared from rat liver, and the activities of catalase, the beta-oxidation system and individual enzymes of beta-oxidation (fatty acyl-CoA oxidase, crotonase, beta-hydroxybutyryl-CoA dehydrogenase and thiolase) were determined. Catalase activity was decreased to about 2% of the control. Among the individual enzymes of the beta-oxidation system, thiolase activity was decreased to 67%, but the activities of fatty acyl-CoA oxidase, crotonase and beta-hydroxybutyryl-CoA dehydrogenase were almost unchanged. The activity of the peroxisomal beta-oxidation system was assayed by measuring palmitoyl-CoA-dependent NADH formation, and the activity of the purified peroxisome preparation was found to be almost unaffected by the administration of aminotriazole. The activity of the system in the aminotriazole-treated preparation was, however, significantly decreased to 55% by addition of 0.1 mM H2O2 to the incubation mixture. Hydrogen peroxide (0.1 mM) reduced the thiolase activity of the aminotriazole-treated peroxisomes to approx. 40%, but did not affect the other activities of the system. Thiolase activity of the control preparation was decreased to 70% by addition of hydrogen peroxide (0.1 mM). The half-life of 0.1 mM H2O2 added to the thiolase assay mixture was 2.8 min in the case of aminotriazole-treated peroxisomes, and 4 s in control peroxisomes. The ultraviolet spectrum of acetoacetyl-CoA (substrate of thiolase) was clearly changed by addition of 0.1 mM H2O2 to the thiolase assay mixture without the enzyme preparation; the absorption bands at around 233 nm (possibly due to the thioester bond of acetoacetyl-CoA) and at around 303 nm (due to formation of the enolate ion) were both significantly decreased. These results suggest that H2O2 accumulated in peroxisomes after aminotriazole treatment may modify both thiolase and its substrate, and consequently suppress the fatty acyl-CoA beta-oxidation. Therefore, catalase may protect thiolase and its substrate, 3-ketoacyl-CoA, by removing H2O2, which is abundantly produced during peroxisomal enzyme reactions.
为了评估过氧化氢酶对过氧化物酶体脂肪酸酰基辅酶Aβ氧化系统的影响，给大鼠施用氨基三唑以抑制其过氧化氢酶活性。在施用氨基三唑2小时后，从大鼠肝脏制备过氧化物酶体，并测定过氧化氢酶、β氧化系统以及β氧化的各个酶（脂肪酰基辅酶A氧化酶、巴豆酸酶、β-羟基丁酰辅酶A脱氢酶和硫解酶）的活性。过氧化氢酶活性降至对照的约2%。在β氧化系统的各个酶中，硫解酶活性降至67%，但脂肪酰基辅酶A氧化酶、巴豆酸酶和β-羟基丁酰辅酶A脱氢酶的活性几乎未变。通过测量棕榈酰辅酶A依赖性NADH的形成来测定过氧化物酶体β氧化系统的活性，发现纯化的过氧化物酶体制剂的活性几乎不受氨基三唑施用的影响。然而，在孵育混合物中加入0.1 mM H2O2后，氨基三唑处理制剂中该系统的活性显著降至55%。过氧化氢（0.1 mM）使氨基三唑处理的过氧化物酶体的硫解酶活性降至约40%，但不影响该系统的其他活性。在对照制剂中加入过氧化氢（0.1 mM）后，硫解酶活性降至70%。在氨基三唑处理的过氧化物酶体的情况下，添加到硫解酶测定混合物中的0.1 mM H2O2的半衰期为2.8分钟，而在对照过氧化物酶体中为4秒。在没有酶制剂的情况下，向硫解酶测定混合物中加入0.1 mM H2O2会明显改变乙酰乙酰辅酶A（硫解酶的底物）的紫外光谱；233 nm左右的吸收带（可能由于乙酰乙酰辅酶A的硫酯键）和303 nm左右的吸收带（由于烯醇离子的形成）均显著降低。这些结果表明，氨基三唑处理后过氧化物酶体中积累的H2O2可能会修饰硫解酶及其底物，从而抑制脂肪酸酰基辅酶Aβ氧化。因此，过氧化氢酶可能通过去除过氧化物酶体酶反应过程中大量产生的H2O2来保护硫解酶及其底物3-酮酰基辅酶A。
