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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Catalysis of electron transfer during activation of O-2 by the flavoprotein glucose oxidase
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Catalysis of electron transfer during activation of O-2 by the flavoprotein glucose oxidase

机译：黄素蛋白葡萄糖氧化酶激活O-2期间的电子转移催化

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摘要

Two prototropic forms of glucose oxidase undergo aerobic oxidation reactions that convert FADH(-) to FAD and form H2O2 as a product. Limiting rate constants of k(cat)/K-M(O-2) = (5.7 +/- 1.8) x 10(2) M-1.s(-1) and k(cat)/K-M(O-2) = (1.5 +/- 0.3) x 10(6) M-1.s(-1) are observed at high and low pH, respectively. Reactions exhibit oxygen-18 kinetic isotope effects but no solvent kinetic isotope effects, consistent with mechanisms of rate-limiting electron transfer from flavin to O-2. Site-directed mutagenesis studies reveal that the pH dependence of the rates is caused by protonation of a highly conserved histidine in the active site. Temperature studies (283323 K) indicate that protonation of His-516 results in a reduction of the activation energy barrier by 6.0 kcal.mol(-1) (0.26 eV). Within the context of Marcus theory, catalysis of electron transfer is attributed to a 19-kcal.mol(-1) (0.82 eV) decrease in the reorganization energy and a much smaller 2.2-kcal.mol(-1) (0.095 eV) enhancement of the reaction driving force. An explanation is advanced that is based on changes in outer-sphere reorganization as a function of pH. The active site is optimized at low pH, but not at high pH or in the H516A mutant where rates resemble the uncatalyzed reaction in solution. [References: 61]
机译：葡萄糖氧化酶的两种质子传递形式经历需氧氧化反应，将FADH（-）转化为FAD并形成H2O2作为产物。极限速率常数k（cat）/ KM（O-2）=（5.7 +/- 1.8）x 10（2）M-1.s（-1）和k（cat）/ KM（O-2）=在高和低pH下分别观察到（1.5 +/- 0.3）x 10（6）M-1.s（-1）。反应表现出氧18的动力学同位素效应，但没有溶剂的动力学同位素效应，这与速率限制电子从黄素到O-2的转移机理一致。定点诱变研究表明，速率的pH依赖性是由活性位点中高度保守的组氨酸的质子化引起的。温度研究（283323 K）表明，His-516的质子化导致活化能垒降低了6.0 kcal.mol（-1）（0.26 eV）。在马库斯理论的背景下，电子转移的催化作用归因于重组能量降低19-kcal.mol（-1）（0.82 eV）和小得多的2.2-kcal.mol（-1）（0.095 eV）增强反应驱动力。提出了基于外球重组随pH的变化的解释。活性位点在低pH值下优化，但在高pH值下或H516A突变体中最优化，其速率类似于溶液中未催化的反应。 [参考：61]

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