Conversion of 3Fe-4S to 4Fe-4S clusters in native pyruvate formate-lyase activating enzyme: Mossbauer characterization and implications for mechanism
Conversion of 3Fe-4S to 4Fe-4S clusters in native pyruvate formate-lyase activating enzyme: Mossbauer characterization and implications for mechanism
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DOI：
10.1021/ja003335p
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发表时间：
2000-12-20
影响因子：
15
通讯作者：
Broderick, JB
中科院分区：
文献类型：
--
作者：
Krebs, C;Henshaw, TF;Broderick, JB
丙酮酸甲酸酶激活酶利用铁硫簇和s -腺苷蛋氨酸在丙酮酸甲酸裂解酶上产生催化必需的甘酰基自由基。利用变温度（4.2-200 K）和变场（0.05-8 T）穆斯堡尔光谱对厌氧分离丙酮酸甲酸裂解酶激活酶和二硫代酸还原酶中存在的铁-硫团簇进行了表征。对Mossbauer数据的详细分析表明，厌氧分离酶含有Fe-S簇的混合物，其中立方体[3Fe-4S]（+）簇为主要簇形式，占总铁的66%。其他形式包括[2Fe-2S](2+)（占总铁的12%）和[4Fe-4S](2+)（占总铁的8%）。仔细检查在各种应用领域记录的穆斯堡尔光谱，发现第四种光谱成分被分配到线性[3Fe-4S](+)（类似于总Fe的10%）。有趣的是，二硫代硫酸盐还原分离的酶，将所有簇类型转化为2+（占总铁的66%）和1+（占总铁的12%）混合氧化态的[4Fe-4S]形式。根据这些结果讨论了铁硫团簇在自由基生成中的作用。
Pyruvate formate-ly ase activating enzyme utilizes an iron-sulfur cluster and S-adenosylmethionine to generate the catalytically essential glycyl radical on pyruvate formate-lyase. Variable-temperature (4.2-200 K) and variable-field (0.05-8 T) Mossbauer spectroscopy has been used to characterize the iron-sulfur clusters present in anaerobically isolated pyruvate formate-lyase activating enzyme and in the dithionite-reduced form of the enzyme. Detailed analysis of the Mossbauer data indicates that the anaerobically isolated enzyme contains a mixture of Fe-S clusters with the cuboidal [3Fe-4S](+) clusters as the primary cluster form, accounting for 66% of the total iron. Other forms present include [2Fe-2S](2+) (12% of total Fe) and [4Fe-4S](2+) (8% of total iron). Careful examination of Mossbauer spectra recorded at various applied fields reveal a fourth spectral component which is assigned to a linear [3Fe-4S](+) (similar to 10% of total Fe). Reduction of the as-isolated enzyme by dithionite, interestingly, converts all cluster types into the [4Fe-4S] form with a mixture of 2+ (66% of total iron) and 1+ (12% of total iron) oxidation states. These results are discussed in light of the proposed role for the iron-sulfur cluster in radical generation.