title: A deletion mutant lacking three out of four transfer RNA genes upstream of the coding region of tufB.
abstract: Of the two plasmids pTUB1 and pTUB2 constructed by cloning of the 8.9 kb EcoRI fragment carrying tufB (Miyajima, A., Shibuya, M., & Kaziro, Y. (1979) FEBS Lett. 102, 207-210), pTUB2 possesses a deletion of about 0.3 kb. Restriction and sequence analyses have located the deletion in the region of the four tRNA genes thrU-tyrU-glyT-thrT upstream of the tufB structural gene. As a result of homologous recombination between thrU and thrT, the four tRNA genes have been replaced by a single thrU-thrT hybrid gene. The deletion of the three tRNA genes does not significantly alter the in vivo expression of tufB as assessed by the kirromycin-sensitive phenotype of the transformant cells and by the synthesis of EF-Tu in mini-cells. Nor does the deletion affect the synthesis of beta-galactosidase in lysogens carrying a lambda transducing phage with a tufB-lacZ fusion. Transcription of tufB and synthesis of EF-Tu in a cell-free transcription-translation coupled system were essentially the same, regardless of whether pTUB1 or pTUB2 DNA was used as a template. Likewise, 0.2 mM ppGpp inhibits the synthesis of tufB mRNA on both pTUB1 and pTUB2 templates to the same extent. We concluded that the replacement by thrU-thrT hybrid gene of the four tRNA genes upstream of the tufB coding region does not significantly affect either in vivo or in vitro expression of tufB.
mesh_terms: D010582:Bacteriophage lambda / Q000378:metabolism; D001483:Base Sequence; D002474:Cell-Free System; D002872:Chromosome Deletion; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000235:genetics*; D009154:Mutation; D010444:Peptide Elongation Factor Tu; D010445:Peptide Elongation Factors / Q000235:genetics*; D010957:Plasmids; D011728:Pyridones / Q000494:pharmacology; D012329:RNA, Bacterial / Q000235:genetics*; D012343:RNA, Transfer / Q000235:genetics*
chemical_list: D010445:Peptide Elongation Factors; D011728:Pyridones; D012329:RNA, Bacterial; D012343:RNA, Transfer; D010444:Peptide Elongation Factor Tu; C008536:mocimycin