The [4Fe-4S]1+ cluster of pyruvate formate-lyase activating enzyme generates the glycyl radical on pyruvate formate-lyase:: EPR-detected single turnover
The [4Fe-4S]1+ cluster of pyruvate formate-lyase activating enzyme generates the glycyl radical on pyruvate formate-lyase:: EPR-detected single turnover
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DOI：
10.1021/ja002012q
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发表时间：
2000-08-30
影响因子：
15
通讯作者：
Broderick, JB
中科院分区：
文献类型：
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作者：
Henshaw, TF;Cheek, J;Broderick, JB
丙酮酸甲酸裂解酶激活酶（PFL- ae）在PFL上产生催化必需的甘酰基自由基（Scheme 1），是利用铁硫簇和s -腺苷蛋氨酸（AdoMet）作为自由基生成所需辅因子的一类新兴酶的代表成员。这一类包括相关的激活酶，如来自大肠杆菌的厌氧核糖核苷酸还原酶激活酶（aRNR-AE）， 2以及生物素合成酶，3,4硫辛酸合成酶，5,6和赖氨酸氨基互换酶（LAM）。尽管这些酶在功能上各不相同，但它们被认为具有共同的关键机制特征，包括产生中间的5 ' -脱氧腺苷基自由基，该自由基通过氢原子提取启动催化作用。同位素标记为PFL-AE和LAM的这种机制提供了间接证据。最近，Frey和他的同事们的出色工作为LAM的5 ' -脱氧腺苷基的烯丙基类似物提供了直接的光谱证据。围绕这组酶的一个中心问题是铁硫簇参与5 ' -脱氧腺苷基中间体生成的机制。多种铁硫簇，包括[2Fe-2S]，[3Fe-4S]和[4Fe-4S]，已经在这些adomet依赖性酶中被鉴定出来。2- 7,11 -13然而，要明确地确定与催化相关的簇是很困难的。A [4Fe-4S] 1+被认为是含有[4Fe-4S] 1+ EPR信号的aRNR、2a和LAM的活性簇，并被证明具有催化活性。7a我们在这里报道了在限制还原剂条件下的PFL- ae，每个[4Fe-4S] 1+簇能够在PFL上生成单个甘酰基自由基。我们的结果提供了第一个直接的定量光谱证据，证明PFL-AE的[4Fe-4S] 1+是催化相关的簇，并且
Pyruvate formate-lyase activating enzyme (PFL-AE), which generates the catalytically essential glycyl radical on PFL (Scheme 1), 1 is a representative member of an emerging group of enzymes that utilize iron-sulfur clusters and S-adenosylmethionine (AdoMet) as required cofactors in radical generation. This group includes related activating enzymes such as the anaerobic ribonucleotide reductase activating enzyme (aRNR-AE) from E. coli, 2 as well as biotin synthase, 3, 4 lipoic acid synthase, 5, 6 and lysine aminomutase (LAM). 7 Though diverse in function, these enzymes have been proposed to have in common key mechanistic features including the generation of an intermediate 5′-deoxyadenosyl radical that initiates catalysis by hydrogen atom abstraction. Isotopic labeling has provided indirect evidence for such a mechanism for PFL-AE and LAM. 8, 9 Recently, elegant work by Frey and co-workers has provided direct spectroscopic evidence for an allylic analogue of the 5′-deoxyadenosyl radical for LAM. 10A central question surrounding this group of enzymes is the mechanism by which the iron-sulfur clusters participate in generation of the 5′-deoxyadenosyl radical intermediate. A variety of iron-sulfur clusters, including [2Fe-2S],[3Fe-4S], and [4Fe-4S], have been identified in these AdoMet-dependent enzymes. 2-7, 11-13 It has been difficult, however, to identify unequivocally the catalytically relevant cluster. A [4Fe-4S] 1+ has been implicated as the active cluster for aRNR, 2a and LAM containing a [4Fe-4S] 1+ EPR signal has been shown to be catalytically active. 7a We report here that for PFL-AE under conditions of limiting reductant, each [4Fe-4S] 1+ cluster is capable of generating a single glycyl radical on PFL. Our results provide the first direct quantitative spectroscopic evidence that the [4Fe-4S] 1+ of PFL-AE is the catalytically relevant cluster, and