Figure 3.
Amount and stability of RNase R mutant proteins. (A) Amount of wild type (WT) and K544R mutant RNase R protein in exponential phase (Exp) and stationary phase (Sta) cells. (B) Half-life of wild type and K544R mutant RNase R in exponential and stationary phase cells. (C) Amount of wild type and K544A mutant RNase R protein in exponential phase and stationary phase cells. (D) Half-life of K544A mutant RNase R in exponential and stationary phase cells. (E) Amount of wild type and E764A, D766A mutant RNase R protein in exponential phase and stationary phase cells. (F) Half-life of E764A, D766A mutant RNase R in exponential and stationary phase cells. The indicated cells were grown in YT medium, treated with chloramphenicol and assayed for RNase R protein by immunoblotting as described in “Experimental Procedures”. For panels (A) through (F), 5 μg of total protein were added to each lane. (G) Acetylation analysis of RNase R mutant proteins. Equal amounts of purified exponential phase and stationary phase wild type, K544R, K544A and E764A, D766A mutant proteins (1 ng) were resolved on 8% SDS-PAGE and then probed with RNase R antibody and anti acetylated-lysine monoclonal antibody (Ac-K), respectively. Each gel shown in panels (A) through (G) is a representative experiment carried out at least twice.