Common thiolation mechanism in the biosynthesis of tRNA thiouridine and sulphur-containing cofactors.
2-Thioribothymidine (s(2)T), a modified uridine, is found at position 54 in transfer RNAs (tRNAs) from several thermophiles; s(2)T stabilizes the L-shaped structure of tRNA and is essential for growth at higher temperatures. Here, we identified an ATPase (tRNA-two-thiouridine C, TtuC) required for the 2-thiolation of s(2)T in Thermus thermophilus and examined in vitro s(2)T formation by TtuC and previously identified s(2)T-biosynthetic proteins (TtuA, TtuB, and cysteine desulphurases). The C-terminal glycine of TtuB is first activated as an acyl-adenylate by TtuC and then thiocarboxylated by cysteine desulphurases. The sulphur atom of thiocarboxylated TtuB is transferred to tRNA by TtuA. In a ttuC mutant of T. thermophilus, not only s(2)T, but also molybdenum cofactor and thiamin were not synthesized, suggesting that TtuC is shared among these biosynthetic pathways. Furthermore, we found that a TtuB-TtuC thioester was formed in vitro, which was similar to the ubiquitin-E1 thioester, a key intermediate in the ubiquitin system. The results are discussed in relation to the mechanism and evolution of the eukaryotic ubiquitin system.
Shigi
Naoki
N
Biomedicinal Information Research Center (BIRC), National Institute of Advanced Industrial Science and Technology (AIST), Tokyo, Japan.
Sakaguchi
Yuriko
Y
Asai
Shin-Ichi
S
Suzuki
Tsutomu
T
Watanabe
Kimitsuna
K
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D003067:Coenzymes / Q000096:biosynthesis*; D017353:Gene Deletion; D008667:Metalloproteins / Q000096:biosynthesis; D008954:Models, Biological; D000090006:Molybdenum Cofactors; D011621:Pteridines; D012343:RNA, Transfer / Q000378:metabolism*; D016963:Thermus thermophilus / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D013831:Thiamine / Q000096:biosynthesis; D013891:Thiouridine / Q000378:metabolism*
D000251:Adenosine Triphosphatases; D001426:Bacterial Proteins; D003067:Coenzymes; D008667:Metalloproteins; D000090006:Molybdenum Cofactors; D011621:Pteridines; D012343:RNA, Transfer; D013831:Thiamine; D013891:Thiouridine; C022261:molybdopterin