Why Nature Uses Radical SAM Enzymes so Widely: Electron Nuclear Double Resonance Studies of Lysine 2,3-Aminomutase Show the 5'-dAdo• "Free Radical" Is Never Free.
Why Nature Uses Radical SAM Enzymes so Widely: Electron Nuclear Double Resonance Studies of Lysine 2,3-Aminomutase Show the 5'-dAdo• "Free Radical" Is Never Free.
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DOI：
10.1021/jacs.5b00498
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发表时间：
2015-06-10
影响因子：
15
通讯作者：
Hoffman BM
中科院分区：
文献类型：
--
作者：
Horitani M;Byer AS;Shisler KA;Chandra T;Broderick JB;Hoffman BM
赖氨酸2，3-氨基变位酶（LAM）是一种自由基S-腺苷-L-甲硫氨酸（SAM）酶，并且与该超家族的其他成员一样，LAM利用自由基产生机制，包括通过经典的五元N，O螯合环锚定到[4Fe-4S]簇的独特Fe的SAM。催化作用由SAM S-C5′键的还原裂解引发，产生高度反应性的5′-脱氧腺苷自由基（5′-dAdo·），与B12自由基酶中均裂Co-C键裂解产生的自由基相同。SAM替代物S-3′，4 ′-脱水腺苷-L-甲硫氨酸（anSAM）可通过稳定的烯丙基脱水腺苷自由基（anAdo·）替代SAM作为辅因子参与LAM将L-α-赖氨酸异构化为L-β-赖氨酸。在这里，在13 C、2 H和15 N标记的赖氨酸存在下，anAdo·自由基的电子核双共振（ENDOR）光谱完成了来自Clostridium subterminale SB 4的LAM的活性位点如何“驯服”5′-dAdo·自由基，防止其进行有害的副反应的图片：LAM中的这种“自由基”永远不会自由。产生自由基的[4 Fe-4S]/SAM结构的低空间需求允许底物靶与S-C5 '键相邻结合，从而使通过该键断裂产生的5'-dAdo·自由基通过向靶进行约0.6次运动和总体约1.5次运动与其配偶体反应，这些运动由与其配偶体的紧密货车范德华接触控制。我们认为，在自由基SAM酶超家族中，对底物的可及性和对反应性C5′自由基的现成控制，以及对整个催化循环中的小运动的“货车德瓦尔斯控制”是常见的，并且是这些酶是自然界中引发自由基反应的优选手段的主要原因。
Lysine 2,3-aminomutase (LAM) is a radical S-adenosyl-L-methionine (SAM) enzyme and, like other members of this superfamily, LAM utilizes radical-generating machinery comprising SAM anchored to the unique Fe of a [4Fe-4S] cluster via a classical five-membered N,O chelate ring. Catalysis is initiated by reductive cleavage of the SAM S–C5′ bond, which creates the highly reactive 5′-deoxyadenosyl radical (5′-dAdo•), the same radical generated by homolytic Co–C bond cleavage in B12 radical enzymes. The SAM surrogate S-3′,4′-anhydroadenosyl-L-methionine (anSAM) can replace SAM as a cofactor in the isomerization of L-α-lysine to L-β-lysine by LAM, via the stable allylic anhydroadenosyl radical (anAdo•). Here electron nuclear double resonance (ENDOR) spectroscopy of the anAdo• radical in the presence of 13C, 2H, and 15N-labeled lysine completes the picture of how the active site of LAM from Clostridium subterminale SB4 “tames” the 5′-dAdo• radical, preventing it from carrying out harmful side reactions: this “free radical” in LAM is never free. The low steric demands of the radical-generating [4Fe-4S]/SAM construct allow the substrate target to bind adjacent to the S–C5′ bond, thereby enabling the 5′-dAdo• radical created by cleavage of this bond to react with its partners by undergoing small motions, ~0.6 Å toward the target and ~1.5 Å overall, that are controlled by tight van der Waals contact with its partners. We suggest that the accessibility to substrate and ready control of the reactive C5′ radical, with “van der Waals control” of small motions throughout the catalytic cycle, is common within the radical SAM enzyme superfamily and is a major reason why these enzymes are the preferred means of initiating radical reactions in nature.