The thiolation of uridine 34 in tRNA, which controls protein translation, depends on a [4Fe-4S] cluster in the archaeum Methanococcus maripaludis

• Published in: Scientific reports Vol. 13; no. 1; p. 5351
• Main Authors: Bimai, Ornella, Legrand, Pierre, Ravanat, Jean-Luc, Touati, Nadia, Zhou, Jingjing, He, Nisha, Lénon, Marine, Barras, Frédéric, Fontecave, Marc, Golinelli-Pimpaneau, Béatrice
• Format: Journal Article
• Language: English
• Published: England Nature Publishing Group 01.04.2023; Nature Publishing Group UK; Nature Portfolio
• Subjects: Biochemistry; Biochemistry, Molecular Biology; Catalysis; Crystal structure; Cysteine - metabolism; Cytosol; Enzymes; Humans; Iron-Sulfur Proteins - metabolism; Life Sciences; Methanococcus - genetics; Methanococcus maripaludis; Microbiology and Parasitology; Protein Biosynthesis; RNA, Transfer - genetics; Structural Biology; Sulfur; Sulfur - metabolism; Transfer RNA; Translation; tRNA; Uridine; Uridine - metabolism
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-023-32423-9

Thiolation of uridine 34 in the anticodon loop of several tRNAs is conserved in the three domains of life and guarantees fidelity of protein translation. U34-tRNA thiolation is catalyzed by a complex of two proteins in the eukaryotic cytosol (named Ctu1/Ctu2 in humans), but by a single NcsA enzyme in archaea. We report here spectroscopic and biochemical experiments showing that NcsA from Methanococcus maripaludis (MmNcsA) is a dimer that binds a [4Fe-4S] cluster, which is required for catalysis. Moreover, the crystal structure of MmNcsA at 2.8 Å resolution shows that the [4Fe-4S] cluster is coordinated by three conserved cysteines only, in each monomer. Extra electron density on the fourth nonprotein-bonded iron most likely locates the binding site for a hydrogenosulfide ligand, in agreement with the [4Fe-4S] cluster being used to bind and activate the sulfur atom of the sulfur donor. Comparison of the crystal structure of MmNcsA with the AlphaFold model of the human Ctu1/Ctu2 complex shows a very close superposition of the catalytic site residues, including the cysteines that coordinate the [4Fe-4S] cluster in MmNcsA. We thus propose that the same mechanism for U34-tRNA thiolation, mediated by a [4Fe-4S]-dependent enzyme, operates in archaea and eukaryotes.