Radical S‑Adenosyl‑l‑Methionine Enzyme PylB: A C‑Centered Radical to Convert l‑Lysine into (3R)‑3-Methyl‑d‑Ornithine

• Published in: Journal of the American Chemical Society 2024-03, Vol.146 (10), p.6493-6505
• Main Authors: Soualmia, Feryel, Cherrier, Mickael V., Chauviré, Timothée, Mauger, Mickaël, Tatham, Philip, Guillot, Alain, Guinchard, Xavier, Martin, Lydie, Amara, Patricia, Mouesca, Jean-Marie, Daghmoum, Meriem, Benjdia, Alhosna, Gambarelli, Serge, Berteau, Olivier, Nicolet, Yvain
• Format: Article
• Language: English
• Subjects: Biochemistry; Biochemistry, Molecular Biology; Catalysis; Chemical Sciences; Electron Spin Resonance Spectroscopy; Life Sciences; Lysine; Methionine; or physical chemistry; Ornithine - analogs & derivatives; Racemethionine; S-Adenosylmethionine - metabolism; Theoretical and
• ISSN: 0002-7863; 1520-5126; 1520-5126
• DOI: 10.1021/jacs.3c03747

PylB is a radical S-adenosyl-l-methionine (SAM) enzyme predicted to convert l-lysine into (3R)-3-methyl-d-ornithine, a precursor in the biosynthesis of the 22nd proteogenic amino acid pyrrolysine. This protein highly resembles that of the radical SAM tyrosine and tryptophan lyases, which activate their substrate by abstracting a H atom from the amino-nitrogen position. Here, combining in vitro assays, analytical methods, electron paramagnetic resonance spectroscopy, and theoretical methods, we demonstrated that instead, PylB activates its substrate by abstracting a H atom from the Cγ position of l-lysine to afford the radical-based β-scission. Strikingly, we also showed that PylB catalyzes the reverse reaction, converting (3R)-3-methyl-d-ornithine into l-lysine and using catalytic amounts of the 5′-deoxyadenosyl radical. Finally, we identified significant in vitro production of 5′-thioadenosine, an unexpected shunt product that we propose to result from the quenching of the 5′-deoxyadenosyl radical species by the nearby [Fe4S4] cluster.