Identification of Two tRNA Thiolation Genes Required for Cell Growth at Extremely High Temperatures.

Thermostability of tRNA in thermophilic bacteria is effected by post-transcriptional modifications, such as 2-thioribothymidine (s²T) at position 54. Using a proteomics approach, we identified two genes (ttuA and ttuB; tRNA-two-thiouridine) that are essential for the synthesis of s²T in Thermus thermophilus. Mutation of either gene completely abolishes thio-modification of s²T, and these mutants exhibit a temperature-sensitive phenotype. These results suggest that bacterial growth at higher temperatures is achieved through the thermal stabilization of tRNA by a 2-thiolation modification. TtuA (TTC0106) is possibly an ATPase possessing a P-loop motif. TtuB (TTC0105) is a putative thio-carrier protein that exhibits significant sequence homology with ThiS of the thiamine synthesis pathway. Both TtuA and TtuB are required for in vitro s²T formation in the presence of cysteine and ATP. The addition of cysteine desulfurases such as IscS (TTC0087) or SufS (TTC1373) enhances the sulfur transfer reaction in vitro. [ABSTRACT FROM AUTHOR]