What strategies are employed by organisms that can survive under high-temperature environments such as hydrothermal vents? One of the answers is a post-transcriptional modification of transfer ribonucleic acid (tRNA). tRNA transfers amino acids according to its anticodon for protein synthesis. However, immature tRNA cannot function without modifications due to the low thermal stability and translational fidelity. Our target, 5-methyl-2-thiouridine modification at position 54 of tRNA (thiolation, m5s2U54) improves the melting temperature of tRNA from 65°C to 89°C, which is an essential modification for hyperthermophiles to survive above 70°C.
We have successfully determined the crystal structure of 2-thiouridine synthetase (TtuA) catalyzing the abovementioned tRNA thiolation, complexed with sulfur donor protein TtuB. Furthermore, we revealed that an oxygen-sensitive co-factor [4Fe-4S] cluster is essential for the enzymatic activity of TtuA[1]. We also found that one exposed iron (the unique iron) of the [4Fe-4S] that is not bound to the cysteine residues of TtuA receives the sulfur of TtuB[2].
Recently, it is reported that an abnormal thiolation at position 34 of human tRNA (position 1 of anticodon) is associated with cancer invasion/metastasis and mitochondrial disease. Interestingly, the structure and function of Ncs6 catalyzing this thiolation is similar to that of TtuA, but it has been suggested that the type of iron-sulfur cluster may be different. It is also unclear how TtuA/Ncs6 interacts with tRNA and activates tRNA. To address these problems, we aim to elucidate the reaction mechanism of tRNA thiolation.
Reference
- M. Chen, et al., PNAS 114, 4954–4959 (2017). https://www.pnas.org/doi/10.1073/pnas.1615585114
- M. Chen, et al., Commun. Biol. 3, 168 (2020). https://www.nature.com/articles/s42003-020-0895-3