Suppression of thermosensitive peptidyl-tRNA hydrolase mutation in Escherichia coli by gene duplication.
Peptidyl-tRNA hydrolase (Pth) in Escherichia coli is required to recycle tRNA molecules that dissociate from the ribosome as peptidyl-tRNA during protein synthesis. At non-permissive temperatures, strains with a thermosensitive mutation affecting the enzyme accumulate peptidyl-tRNA, cease protein synthesis and die. The rate of reversion of this mutation to thermoresistance varies widely according to the genetic background of the cell and the temperature of selection; under certain conditions, reversion can occur at rates approaching 10(-3) per cell per generation. In such revertants, a chromosomal pth gene can be replaced by an inactivated gene, restoring thermosensitive growth in most cases. PCR amplification experiments and Southern blots show the presence of both normal and inactivated copies of the gene, demonstrating that gene duplication has occurred in the revertants. Estimation of intracellular peptidyl-tRNA hydrolase by Western blotting confirms this explanation of the mechanism of high-frequency reversion to thermoresistance.
Menez
Jeanne
J
UPR9073 du CNRS, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, F-75005 Paris, France1.
Remy
Elisabeth
E
Aventis Pharma SA, 13 Quai Jules Guesde, 94403 Vitry sur Seine Cedex, France2.
Buckingham
Richard H
RH
UPR9073 du CNRS, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, F-75005 Paris, France1.
D015139:Blotting, Southern; D015153:Blotting, Western; D002265:Carboxylic Ester Hydrolases / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D020440:Gene Duplication; D006361:Heating; D016133:Polymerase Chain Reaction; D013489:Suppression, Genetic
D002265:Carboxylic Ester Hydrolases; C019674:aminoacyl-tRNA hydrolase