A novel multistep mechanism for oxygen binding to ferrous hemoproteins: Rapid kinetic analysis of ferrous-dioxy myeloperoxidase (compound III) formation
A novel multistep mechanism for oxygen binding to ferrous hemoproteins: Rapid kinetic analysis of ferrous-dioxy myeloperoxidase (compound III) formation
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DOI：
10.1021/bi049541h
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发表时间：
2004-09-14
期刊：
影响因子：
2.9
通讯作者：
Hazen, SL
中科院分区：
文献类型：
--
作者：
Abu-Soud, HM;Raushel, FA;Hazen, SL
髓过氧化物酶（MPO）是一种血红素蛋白，它在氧化反应的催化中使用H2 O2作为电子受体，被认为是炎症损伤和心血管疾病的参与者。一旦释放，关闭这种酶的机制，防止不必要的组织损伤，知之甚少。我们最近证明，MPO血红素减少导致血红素口袋的崩溃，如双原子配体结合血红素铁的速率显着减少所监测的。使用光谱和快速动力学测量，我们现在证明，分子氧（O-2）结合到铁MPO（MPO-Fe（II））在一个独特的和新颖的机制。而不是发生通过一个简单的，可逆的，一步的机制，是典型的O-2结合到其他亚铁血红素蛋白，反应涉及几个动力学和光谱可区分的中间体。二极管阵列分光光度法和停流的研究表明，形成的MPO-Fe（II）-O-2复合物包括至少三个基本步骤，并包括至少两个顺序的瞬态中间体。第一步涉及可逆形成的瞬态中间体通过O-2-依赖的机制，其次是两个连续的O-2-独立的步骤，似乎是构象的起源。深入了解MPO的失活机制和O-2与血红素蛋白结合的新模式可能为理解MPO的催化作用提供重要线索。
Myeloperoxidase (MPO), a hemoprotein that uses H2O2 as the electron acceptor in the catalysis of oxidative reactions, is implicated as a participant in inflammatory injury and cardiovascular diseases. Mechanisms for turning off this enzyme once released, preventing unwanted tissue injury, are poorly understood. We recently demonstrated that MPO heme reduction causes collapse of the heme pocket, as monitored by significant reductions in the rates of diatomic ligand binding to the heme iron. Using spectral and rapid kinetic measurements, we now demonstrate that molecular oxygen (O-2) binds to ferrous MPO (MPO-Fe(II)) in a distinct and novel mechanism. Rather than occurring through a simple, reversible, one-step mechanism, as is typical for O-2 binding to other ferrous hemoproteins, the reaction involves several kinetically and spectroscopically distinguishable intermediates. Diode array spectrophotometric and stopped-flow studies reveal that the formation of the MPO-Fe(II)-O-2 complex consists of at least three elementary steps and includes at least two sequential transient intermediates. The first step involves reversible formation of a transient intermediate via an O-2-dependent mechanism, followed by two sequential O-2-independent steps that appear to be conformational in origin. Insights into mechanisms for inactivating MPO and the novel mode Of O-2 binding to the hemoprotein may provide important clues toward understanding the catalytic action of MPO.