X-ray spectroscopy of the iron site in soybean lipoxygenase-1: changes in coordination upon oxidation or addition of methanol.
X-ray spectroscopy of the iron site in soybean lipoxygenase-1: changes in coordination upon oxidation or addition of methanol.
复制标题
大豆脂氧合酶-1 中铁位点的 X 射线光谱：氧化或添加甲醇时配位的变化。
DOI：
10.1021/bi00254a011
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发表时间：
1994
期刊：
影响因子：
2.9
通讯作者：
Nelson,MJ
中科院分区：
文献类型：
--
作者：
Scarrow,RC;Trimitsis,MG;Buck,CP;Grove,GN;Cowling,RA;Nelson,MJ
摘要：利用铁K边X射线能谱（XANES和EXAFS）研究了大豆脂氧合酶-1（SLO）冷冻溶液中的铁配位。天然铁（II）脂氧合酶的Is-3D前边缘过渡的强度大于六配位高自旋铁（II）模型复合物，但与五配位模型的强度相当。这一点和EXAFS测定的相对较短的平均键长（2.13 A）表明，我们冷冻样品中的天然脂氧合酶是五配位的，排除了可能的键长超过2.5 A。铁（II）天然脂氧合酶的协调变化时，甲醇（低至0.1%）或甘油（20%）添加到缓冲液冷冻前。甲醇的加入减少了前边缘过渡，并增加了EXAFS衍生的键长0.04 A，表明六配位的变化。活性铁（III）脂氧合酶的小前边缘功能表明六协调。EXAFS表明一个短的，1.88 A的Fe-O键，其中，考虑到其他光谱和晶体学证据，被分配到配位的氢氧化物。剩余键长的平均值为2.11 A。铁（III）脂氧合酶中的铁配位受醇的存在的影响小于铁（II）酶中的位点。键价总和表明，来自我们的EXAFS分析的脂氧合酶的键长是可比的晶体学特征的模型复合物。SLON（天然SLO）中配位数的灵活性和SLOA（活性SLO）中[FemOH] 2+单元的存在可能具有重要的机理性。大豆脂氧合酶-1是催化多不饱和脂肪酸氧化的非血红素铁酶（Gardner，1991; Siedow，1991）。例如，它催化亚油酸与分子氧反应生成13-HPOD。1在各种条件下由脂氧合酶产生的烷基和过氧基脂肪酸自由基的观察（Chamulitrat & Mason，1989;纳尔逊& Cowling，1990;纳尔逊et al.，1990，1994）支持活性位点铁离子氧化1，4-二烯的机制，
Revised Manuscript Received October 14, 1994% abstract: Iron K-edge X-ray spectroscopy (XANES and EXAFS) was used to study iron coordination in frozen solutions of soybean lipoxygenase-1 (SLO). The intensity of the Is—3d pre-edge transition of native iron (II) lipoxygenase is greater than what was found for six-coordinate high-spin iron (II) model complexes, but comparable to that of a five-coordinate model. This and a relatively short average bond length determined by EXAFS (2.13 A) indicate that the native lipoxygenase in our frozen samples is five-coordinate, excluding possiblebonds longer than 2.5 A. The coordination of the iron (II) innative lipoxygenase changes when methanol (as low as 0.1%) or glycerol (20%) is added to the buffer prior to freezing. The addition of methanol diminishes the pre-edge transition and increases EXAFS-derived bond lengths by 0.04 A, indicating a change to six-coordination. The small pre-edge feature in active iron (III) lipoxygenase suggests six-coordination. EXAFS indicates a short, 1.88 A Fe—O bond, which, given other spectroscopic and crystallographic evidence, is assigned to coordinated hydroxide. The average of the remaining bond lengths is 2.11 A. The iron coordination in iron (III) lipoxygenase is less affected by the presence of alcohols than is the site in the iron (II) enzyme. Bond valence sums indicate that the bond lengths for lipoxygenase derived from our EXAFS analyses are comparable to those of crystallographically characterized model complexes. The flexibility of the coordination number in SLON (native SLO) and the presence of an [FemOH] 2+ unit in SLOA (active SLO) are of possible mechanistic importance.Soybean lipoxygenase-1 is a non-heme iron enzyme that catalyzes the oxygenation of polyunsaturated fatty acids (Gardner, 1991; Siedow, 1991). For example, it catalyzes the reaction of linoleic acid with dioxygen to produce 13-HPOD. 1 The observation of alkyl and peroxyl fatty acid radicals generated by lipoxygenase under a variety of conditions (Chamulitrat & Mason, 1989; Nelson & Cowling, 1990; Nelson et al., 1990, 1994) supports a mechanism in which the active-site ferric ion oxidizes the 1, 4-diene of