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首页> 外文期刊>The biochemical journal >Kinetic study of the inactivation of ascorbate peroxidase by hydrogen peroxide
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Kinetic study of the inactivation of ascorbate peroxidase by hydrogen peroxide

机译：过氧化氢抗坏血酸过氧化物酶的动力学研究

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pThe activity of ascorbate peroxidase (APX) has been studied with Hsub2/subOsub2/sub and various reducing substrates. The activity decreased in the order pyrogallol & ascorbate & guaiacol & 2,2?-azino-bis-(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS). The inactivation of APX with Hsub2/subOsub2/sub as the sole substrate was studied. The number of Hsub2/subOsub2/sub molecules required for maximal inactivation of the enzyme was determined as approx. 2.5. Enzymic activity of approx. 20% of the original remained at the end of the inactivation process (i.e. approx. 20% resistance) when ascorbate or ABTS was used as the substrate in activity assays. With pyrogallol or guaiacol no resistance was seen. Inactivation by Hsub2/subOsub2/sub followed over time with ascorbate or pyrogallol assays exhibited single-exponential decreases in enzymic activity. Hyperbolic saturation kinetics were observed in both assay systems; a similar dissociation constant (0.8 iμ/iM) for Hsub2/subOsub2/sub was obtained in each case. However, the maximum rate constant (λsubmax/sub) obtained from the plots differed depending on the assay substrate. The presence of reducing substrate in addition to Hsub2/subOsub2/sub partly or completely protected the enzyme from inactivation, depending on how many molar equivalents of reducing substrate were added. An oxygen electrode system has been used to confirm that APX does not exhibit a catalase-like oxygen-releasing reaction. A kinetic model was developed to interpret the experimental results; both the results and the model are compared and contrasted with previously obtained results for horseradish peroxidase C. The kinetic model has led us to the conclusion that the inactivation of APX by Hsub2/subOsub2/sub represents an unusual situation in which no enzyme turnover occurs but there is a partition of the enzyme between two forms, one inactive and the other with activity towards reducing substrates such as ascorbate and ABTS only. The partition ratio is less than 1./p
机译：>已经用H 2 O 2 和各种还原基材研究了抗坏血酸过氧化物酶（APX）的活性。订单Pyrogallol＆GT的活动减少;抗坏血酸＆ Guaiacol＆ 2,2？-Azino-BI-（3-乙基苯并噻唑啉-6-磺酸）（ABTS）。研究了APX与H 2 O 2 作为唯一基板的灭活。最大灭活酶所需的H 2 o 2 分子的数量确定为约。 2.5。酶活性约为。当使用抗坏血酸或ABTS作为活性测定中的基材时，20％的原始原始原始原件仍然在灭活过程（即20％抗性）的末端。用吡羟基醇或愈缩蛋白没有抵抗。通过H 2 O 2 随后与抗坏血酸或吡羟吡咯醇测定的时间随之而来的是在酶活性中表现出单指数降低。在两个测定系统中观察到双曲饱和动力学;在每种情况下，获得H 2 O 2 的类似解离常数（0.8℃）。然而，从曲线图获得的最大速率常数（λ<亚>最大）根据测定衬底而不同。除了H 2 O 2-2 之外还存在还原底物的存在，或者部分地或完全保护酶免于灭活，这取决于加入多少还原基质的摩尔当量。氧电极系统已用于确认APX不表现出脱脂酶样氧释放反应。开发了动力学模型以解释实验结果;结果和模型的结果和模型都与先前获得的辣根过氧化物酶C的结果对比。动力学模型导致我们结论是H 2 O 2的灭活APX的灭活代表了不寻常的情况，其中不会发生酶转化率，但是在两个形式之间存在酶的分区，其中一个非活动和另一个，具有仅减少诸如抗坏血酸和ABT的基材的活动。分区比小于1。

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