Fig. 5.

Pulse chase of TraR point mutants in E. coli. Cells expressing wild type or TraR point mutants from a phage T7 promoter were treated with rifampicin to block host transcription, then treated with [³⁵S]methionine for 1 min, followed by addition of excess nonlabeled methionine. OOHL was provided prior to addition of radiolabel. At the indicated intervals after the addition of nonlabeled methionine, aliquots were frozen at −80°C to terminate proteolysis, then thawed and size-fractionated by SDS-PAGE. Radiolabel was quantitated using a Storm PhosphorImager. Calculated TraR half-lives are indicated at the right of each panel and represent the average of two independent experiments.