Mauger Mickaël, Guillot Alain, Seif-Eddine Maryam, Grimaldi Stéphane, Benjdia Alhosna, Berteau Olivier
Université Paris-Saclay, INRAE, AgroParisTech, Micalis Institute, ChemSyBio, 78350 Jouy-en-Josas, France.
Aix Marseille Université, CNRS, IM2B, IMM, BIP UMR 7281, 13009 Marseille, France.
J Am Chem Soc. 2025 Jun 18;147(24):20381-20393. doi: 10.1021/jacs.5c01006. Epub 2025 Jun 9.
Radical SAM enzymes are the most widespread biocatalysts. These metalloenzymes, using -adenosyl-l-methionine (SAM) and a [4Fe-4S] cluster as central cofactors, catalyze a broad range of chemically challenging transformations. The vast majority of radical SAM enzymes initiate their reaction by the homolytic cleavage of the SAM C5'-S bond and the generation of the central 5'-deoxyadenosyl radical (5'-dA·). In this study, by combining spectroscopic approaches with labeling and biochemical analyses, we show that ArsL, the key enzyme in the biosynthesis of the arsenic-containing antibiotic arsinothricin, catalyzes a unique reaction: the addition of the 3-amino-3-carboxypropyl radical (ACP·) to As(III). Remarkably, by exploiting several radical trapping strategies, we demonstrate that in sharp contrast to canonical radical SAM enzymes ArsL cleaves the SAM Cγ-S bond. In addition, using electron paramagnetic resonance (EPR) and hyperfine sublevel correlation (HYSCORE) spectroscopies, we establish that ArsL has a unique SAM binding mode, consistent with its catalytic properties and predicted structure. Notably, EPR and HYSCORE analyses support that SAM interacts with the radical SAM [4Fe-4S] cluster in an uncharacteristic conformation to form ACP·. Collectively, our study reveals that members of the superfamily of radical SAM enzymes are able to finely tune the binding of the SAM cofactor in order to perform unique chemistries.
自由基S-腺苷甲硫氨酸（SAM）酶是分布最为广泛的生物催化剂。这些金属酶以S-腺苷-L-甲硫氨酸（SAM）和一个[4Fe-4S]簇作为中心辅助因子，催化多种具有化学挑战性的转化反应。绝大多数自由基SAM酶通过SAM C5'-S键的均裂以及中心5'-脱氧腺苷自由基（5'-dA·）的生成来启动反应。在本研究中，通过将光谱学方法与标记和生化分析相结合，我们发现含砷抗生素抗霉素生物合成中的关键酶ArsL催化一种独特的反应：3-氨基-3-羧丙基自由基（ACP·）加成到As(III)上。值得注意的是，通过采用多种自由基捕获策略，我们证明与典型的自由基SAM酶形成鲜明对比的是，ArsL断裂的是SAM Cγ-S键。此外，利用电子顺磁共振（EPR）和超精细能级相关（HYSCORE）光谱学，我们确定ArsL具有独特的SAM结合模式，这与其催化特性和预测结构一致。值得注意的是，EPR和HYSCORE分析支持SAM以一种非典型构象与自由基SAM [4Fe-4S]簇相互作用以形成ACP·。总的来说，我们的研究表明自由基SAM酶超家族的成员能够精细调节SAM辅助因子的结合，以便进行独特的化学反应。
