Department of Chemistry and Biochemistry , Montana State University , Bozeman , Montana 59717 , United States.
Department of Chemistry , Northwestern University , Evanston , Illinois 60208 , United States.
Acc Chem Res. 2018 Nov 20;51(11):2611-2619. doi: 10.1021/acs.accounts.8b00356. Epub 2018 Oct 15.
The seeds for recognition of the vast superfamily of radical S-adenosyl-l-methionine (SAM) enzymes were sown in the 1960s, when Joachim Knappe found that the dissimilation of pyruvate was dependent on SAM and Fe(II), and Barker and co-workers made similar observations for lysine 2,3-aminomutase. These intriguing observations, coupled with the evidence that SAM and Fe were cofactors in radical catalysis by these enzyme systems, drew us in the 1990s to explore how Fe(II) and SAM initiate radical reactions. Our early work focused on the same enzyme Knappe had originally characterized: the pyruvate formate-lyase activating enzyme (PFL-AE). Our discovery of an iron-sulfur cluster in this enzyme, together with similar findings for other SAM-dependent enzymes at the time, led to the recognition of an emerging class of enzymes that use iron-sulfur clusters to cleave SAM, liberating the 5'-deoxyadenosyl radical (5'-dAdo•) that initiates radical reactions. A major bioinformatics study by Heidi Sofia and co-workers identified the enzyme superfamily denoted Radical SAM, now known to span all kingdoms of life with more than 100,000 unique sequences encoding enzymes that catalyze remarkably diverse reactions. Despite the limited sequence similarity and vastly divergent reactions catalyzed, the radical SAM enzymes appear to employ a common mechanism for initiation of radical chemistry, a mechanism we have helped to clarify over the last 25 years. A reduced [4Fe-4S] cluster provides the electron needed for the reductive cleavage of SAM. The resulting [4Fe-4S] cluster can be rereduced either by an external reductant, with SAM acting as a cosubstrate, or by an electron provided during the reformation of SAM in cases where SAM is used as a cofactor. The amino and carboxylate groups of SAM bind to the unique iron of the catalytic [4Fe-4S] cluster, placing the sulfonium of SAM in close proximity to the cluster. Surprising recent results have shown that the initiating enzymatic cleavage of SAM generates an organometallic intermediate prior to liberation of 5'-dAdo•, which initiates radical chemistry on the substrate. This organometallic intermediate, denoted Ω, has a 5'-deoxyadenosyl moiety directly bound to the unique iron of the [4Fe-4S] cluster via the 5'-C, giving a structure that is directly analogous to the Co-(5'-C) bond of the organometallic cofactor adenosylcobalamin. Our observation that this intermediate Ω is formed throughout the superfamily suggests that this is a key intermediate in initiating radical SAM reactions, and that organometallic chemistry is much more broadly relevant in biology than previously thought.
对庞大的自由基 S-腺苷甲硫氨酸（SAM）酶超家族的认识始于 20 世纪 60 年代，当时 Joachim Knappe 发现丙酮酸的异化作用依赖于 SAM 和 Fe(II)，Barker 及其同事也对赖氨酸 2,3-氨基转移酶进行了类似的观察。这些有趣的观察结果，加上证据表明 SAM 和 Fe 是这些酶系统中自由基催化的辅助因子，促使我们在 20 世纪 90 年代探索 Fe(II)和 SAM 如何引发自由基反应。我们的早期工作集中在 Knappe 最初表征的同一种酶上：丙酮酸甲酸裂解酶激活酶（PFL-AE）。我们在该酶中发现了一个铁硫簇，同时也在当时的其他依赖 SAM 的酶中发现了类似的发现，这导致了一类新兴酶的出现，这些酶使用铁硫簇来切割 SAM，释放引发自由基反应的 5'-脱氧腺苷自由基（5'-dAdo•）。Heidi Sofia 及其同事的一项主要生物信息学研究确定了被称为 Radical SAM 的酶超家族，现在已知该家族跨越了所有生命领域，拥有超过 100,000 个独特的编码酶序列，这些酶催化着极其多样化的反应。尽管有限的序列相似性和广泛的催化反应，但 Radical SAM 酶似乎采用了一种共同的机制来引发自由基化学，我们在过去 25 年中帮助澄清了这种机制。一个还原的[4Fe-4S]簇为 SAM 的还原裂解提供了所需的电子。所得的[4Fe-4S]簇可以通过外部还原剂重新还原，其中 SAM 作为辅助底物，或者在 SAM 作为辅助因子重新形成时通过在 SAM 中提供电子还原。SAM 的氨基和羧基基团与催化[4Fe-4S]簇的独特铁结合，使 SAM 的亚砜基与簇紧密相邻。最近令人惊讶的结果表明，SAM 的起始酶促裂解在释放 5'-dAdo•之前产生了一个有机金属中间体，该中间体在底物上引发自由基化学。这个有机金属中间体，称为 Ω，通过 5'-C 直接将 5'-脱氧腺苷部分与[4Fe-4S]簇的独特铁结合，形成的结构直接类似于有机金属辅助因子腺苷钴胺素的 Co-(5'-C)键。我们观察到这个中间体 Ω 在整个超家族中形成，这表明它是引发自由基 SAM 反应的关键中间体，并且有机金属化学在生物学中的相关性比以前认为的要广泛得多。
