Excess of charged tRNA(Lys) maintains low levels of peptidyl-tRNA hydrolase in pth(Ts) mutants at a non-permissive temperature

Cellular changes have been monitored during the suppression, mediated by the overproduction of tRNA(Lys), of thermosensitivity in Escherichia coli strain AA7852 carrying a mutation in peptidyl-tRNA hydrolase (Pth) encoded by the pth(Ts) gene. The presence in AA7852 cells of a plasmid bearing lysV gene helped to maintain low levels of the unstable Pth(Ts) protein and to preserve the viability of the mutant line at 41°C whereas plasmids bearing other tRNA genes were ineffective. At 32°C the excess of tRNA(Lys) did not alter the percentages of the free-, charged- or peptidyl-tRNA(Lys) species compared with those found in strains that did not overproduce tRNA(Lys). At 41°C, however, despite increases in the level of peptidyl-tRNA(Lys), the excess tRNA(Lys) helped to maintain the concentration of charged-tRNA(Lys) at a level comparable with that found in non-overproducer cells grown at a permissive temperature. In addition, the excess tRNA(Lys) at 41°C provoked a reduction in the concentrations of various peptidyl-tRNAs, which normally accumulate in pth(Ts) cells, and a proportional increase in the concentrations of the corresponding aminoacyl-tRNAs. The possible mechanism of rescue due to the overexpression of tRNA(Lys) and the causes of tRNA(Lys) starvation in pth(Ts) strains grown at non-permissive temperatures are considered

Suppression of the Pth(Ts) phenotype mediated by the overproduction of tRNA maintains moderate levels of the Pth(Ts) protein

<p><b>Copyright information:</b></p><p>Taken from "Excess of charged tRNA maintains low levels of peptidyl-tRNA hydrolase in (Ts) mutants at a non-permissive temperature"</p><p>Nucleic Acids Research 2006;34(5):1564-1570.</p><p>Published online 15 Mar 2006</p><p>PMCID:PMC1408313.</p><p>© The Author 2006. Published by Oxford University Press. All rights reserved</p> () Depicts the cellular growth of the (Ts) mutant strain AA7852 separately transformed with pVH124 (ΔU, ΔV), pVH125 (U, ΔV) or pVH119 (U, V) incubated at different temperatures. Isolated colonies of the independent transformants were streaked onto LB-Ap plates and incubated overnight at the indicated temperatures. () Presents the immunodetection of Pth(Ts) in the (Ts) mutant strain AA7852 separately transformed with pVH124, pVH125, pVH119, ptRNACCA (X, R, T, M) or pTH2 (W) and grown at 32°C prior to transfer at time = 0 min at 41 or 43°C. The concentration of Pth(Ts) protein was estimated by immunoblot analysis. The left lane shows purified wild-type Pth protein, which migrates slightly faster in SDS–PAGE than the Pth(Ts) variant (arrowed) ()