Department of Physical Chemistry, University of Castilla-La Mancha, Campus Universitario, Albacete, Spain.
Arch Biochem Biophys. 2011 Dec 1;516(1):10-20. doi: 10.1016/j.abb.2011.09.006. Epub 2011 Sep 22.
Hydrogen peroxide triggers a redox cycle between methemoglobin and ferrylhemoglobin, leading to protein inactivation and oxygen evolution. In the present paper, the catalase-like oxygen production by human methemoglobin in the presence of H(2)O(2) was kinetically characterized with a Clark-type electrode. Progress curves showed a pseudo-steady state in the first minutes of the reaction, while double-reciprocal plots were upwardly concave, indicating positive co-operativity dependent upon protein concentration, which is a very unusual kinetic behavior. Addition of superoxide radical scavengers slightly increased activity, suggesting that most oxygen was produced biocatalytically. By considering all the experimental data obtained, a possible mechanism was proposed, including: (a) competition between the one-electron and the two-electron reductions of the oxoferryl free radical species of hemoglobin, giving rise to ferrylhemoglobin and methemoglobin, respectively; (b) competition between the superoxide-dependent inactivation of the protein and its reduction back to the met state. Computer simulations of that model have been performed by numerically integrating the differential equations set describing the mechanism, which was seen to yield predictions of the kinetic parameters variation consistently with the kinetic behavior experimentally observed. We suggest that the catalase-like activity of methemoglobin must predominantly be a biocatalytic reaction that protects the protein against H(2)O(2)-induced suicide inactivation.
过氧化氢在高铁血红蛋白和高铁氧合血红蛋白之间引发氧化还原循环，导致蛋白质失活和氧气释放。在本文中，使用克拉克型电极对人高铁血红蛋白在 H(2)O(2)存在下的过氧化氢酶样产氧进行了动力学表征。进展曲线在反应的最初几分钟内显示出准稳态，而双倒数图向上凹，表明蛋白浓度依赖性的正协同作用，这是一种非常不寻常的动力学行为。添加超氧自由基清除剂后，活性略有增加，表明大部分氧气是生物催化产生的。考虑到获得的所有实验数据，提出了一种可能的机制，包括：(a) 血红蛋白的氧合自由基物种的单电子和双电子还原之间的竞争，分别产生高铁血红蛋白和高铁血红蛋白；(b) 超氧依赖的蛋白质失活与其还原回 Met 态之间的竞争。通过数值积分描述该机制的微分方程集来模拟该模型，发现模型预测的动力学参数变化与实验观察到的动力学行为一致。我们认为，高铁血红蛋白的过氧化氢酶样活性必须主要是一种生物催化反应，可保护蛋白质免受 H(2)O(2)诱导的自杀失活。
