Oxidation and electronic state dependence of proton transfer in the enzymatic cycle of cytochrome P450eryF.
Oxidation and electronic state dependence of proton transfer in the enzymatic cycle of cytochrome P450eryF.
复制标题
细胞色素 P450eryF 酶循环中质子转移的氧化和电子态依赖性。
DOI：
10.1016/s0162-0134(02)00477-4
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发表时间：
2002
影响因子：
3.9
通讯作者：
Harris,DanniL
中科院分区：
文献类型：
--
作者：
Harris,DanniL
氢键网络由极性/酸性氨基酸侧链锚定的氢键水组成，通常存在于 P450 的氧结合裂缝附近。细胞色素 P450eryF(CYP107A1) 的 O2 结合裂口网络模型的密度泛函和量子动力学计算表明，这种结构基序通过双重基态势垒为 7-10 kcal/mol 的多重质子易位机制，促进质子从网络水超快转移到还原含氧亚铁物种的双氧，并且质子转移反应物和受限质子转移产物的能量具有电子和氧化态依赖性[J。是。化学。苏克。 124（2002）1430]。在本研究中，氧化态依赖性的根源在于不同的质子亲和力，而还原的含氧铁血红素的电子态依赖性则源于初始质子转移事件过渡态附近网络拓扑的细微差异。宽松的电势表面扫描和无约束的质子转移产物优化表明，单线态含氧亚铁血红素和四重态还原含氧亚铁血红素物种中的质子转移产物相对于反应物形式不是可行的稳定（结合）态。虽然 H3O+ 的质子亲和力足以使其质子化含氧亚铁和还原的含氧亚铁血红素物质，但氢键网络稳定的水只能质子化还原的含氧亚铁形式。这种解释通过对 P450nor 的 NO 结合还原亚铁血红素的研究得到证实，P450nor 与含氧亚铁血红素是等电子的，并且具有相似的质子亲和力。对更广泛的 O2 结合裂口模型的密度泛函计算支持多质子易位转移机制，但表明还原的含氧亚铁血红素物质的结合双氧上的显着负电荷密度（在其双态基态）充分极化相关的氢键网络，从而产生短、强、低势垒的氢键。计算出的 O-H-O 键距小于 2.55 Å，并且质子转移反应物和初始（突然）质子转移产物具有近简并性。这些低势垒氢键特征，除了发现 1.3 kcal/mol 的（零点未校正）势垒之外，表明质子从水到远端氧气的转移应该是快速、轻松的，并且可能不需要像最初使用较小模型所建议的大曲率隧道。通过 6-脱氧赤酮内酯 B (6-DEB) 模型对还原的含氧亚铁血红素远端氧的质子化进行的初步评估表明它具有低势垒 (3.8 kcal/mol) 和放热性 (−2.9 kcal/mol)。综合结果表明，使用组合的水网络和 6-DEB 底物质子化剂，还原的含氧亚铁血红素的远端氧同时双质子化，导致 O-O 键断裂的可能性。
Hydrogen bond networks, consisting of hydrogen bonded waters anchored by polar/acidic amino acid sidechains, are often present in the vicinity of the oxygen binding clefts of P450s. Density functional and quantum dynamics calculations of a O2binding cleft network model of cytochrome P450eryF(CYP107A1) indicate that such structural motifs facilitate ultrafast proton transfer from network waters to the dioxygen of the reduced oxyferrous species via a multiple proton translocation mechanism with barriers of 7–10 kcal/mol on its doublet ground state, and that the energies of the proton transfer reactant and constrained proton transfer products have an electronic and oxidation state dependence [J. Am. Chem. Soc. 124 (2002) 1430]. In the present study, the origin of the oxidation state dependence is shown to have its roots in differential proton affinities while the electronic state dependence of the reduced oxyferrous heme has its origins in subtle differences in network topologies near the transition state of the initial proton transfer event. Relaxed potential surface scans and unconstrained proton transfer product optimizations indicate that the proton transfer product in both the singlet oxyferrous heme and the reduced oxyferrous heme species in a quartet state are not viable stable (bound) states relative to the reactant form. While the proton affinity of H3O+is sufficient for it to protonate both the oxyferrous and the reduced oxyferrous heme species, hydrogen bond network stabilized water is only capable of protonating the reduced oxyferrous form. This interpretation is substantiated by study of the NO bound reduced ferrous heme of P450nor, which is isoelectronic with the oxyferrous heme and has a similar proton affinity. Density functional calculations on a more extensive O2binding cleft model support the multiple proton translocation mechanism of transfer but indicates that the significant negative charge density on the bound dioxygen of the reduced oxyferrous heme species, in its doublet ground state, polarizes the associated hydrogen bond network sufficiently so as to result in short, strong, low-barrier hydrogen bonds. The computed O–H–O bond distances are less than 2.55 Å and have a near degeneracy of the proton transfer reactant and initial (sudden) proton transfer products. These low-barrier hydrogen bond features, in addition to the finding of a (zero point uncorrected) barrier of 1.3 kcal/mol, indicate that proton transfer from water to the distal oxygen should be rapid, facile and may not require large curvature tunneling as originally suggested by use of a smaller model. An initial assessment of protonation of the reduced oxyferrous heme distal oxygen by a model of 6-deoxyerythronolide B (6-DEB) indicates it to be low barrier (3.8 kcal/mol) and exothermic (−2.9 kcal/mol). The combined results indicate the plausibility of simultaneous diprotonation of the distal oxygen of the reduced oxyferrous heme, leading to O–O bond scission, using the combined water network and 6-DEB substrate protonation agents.