Acetylation Regulates the Stability of a Bacterial Protein: Growth Stage-Dependent Modification of RNase R

Abstract

SUMMARY RNase R, an Escherichia coli exoribonuclease important for degradation of structured RNAs, increases 3- to 10-fold under certain stress conditions due to an increased half-life for this usually unstable protein. Components of the trans-translation machinery, tmRNA and its associated protein, SmpB, are essential for RNase R instability. However, it is not understood why exponential phase RNase R is unstable or how it becomes stabilized in stationary phase. We show here that these phenomena are regulated by acetylation catalyzed by YfiQ protein. One residue, Lys544, is acetylated in exponential phase RNase R, but not in the stationary phase protein, resulting in tighter binding of tmRNA-SmpB to the C-terminal region of exponential phase RNase R, and subsequent proteolytic degradation. Removal of the positive charge at Lys544 or a negative charge in the C-terminal region likely disrupts their interaction facilitating tmRNA-SmpB binding. These findings indicate that acetylation can regulate the stability of a bacterial protein.

Transfer-messenger RNA-SmpB Protein Regulates Ribonuclease R Turnover by Promoting Binding of HslUV and Lon Proteases

“…Protease inhibitor mixture was purchased from Calbiochem. Purified tmRNA, His-and GST-tagged SmpB and RNase R were described previously (16,17). All other materials were reagent grade.…”

“…Recent work from our laboratory has greatly increased our understanding of the mechanism of this regulation (15)(16)(17)(18). RNase R is an extremely unstable protein in exponential phase cells with a half-life of ϳ10 min (15), whereas it is stabilized under stress conditions leading to its relative elevation (13)(14)(15).…”

“…Elimination of either of these factors or deletion of the C-terminal region stabilizes RNase R in exponential phase cells (16). Mass spectrometric analysis of purified RNase R demonstrated that exponential phase RNase R is post-translationally modified by acetylation of a single residue, Lys 544 (17), a reaction catalyzed by protein lysine acetyltransferase (17,18). In contrast, RNase R is unmodified in stationary phase cells (17).…”

“…Mass spectrometric analysis of purified RNase R demonstrated that exponential phase RNase R is post-translationally modified by acetylation of a single residue, Lys 544 (17), a reaction catalyzed by protein lysine acetyltransferase (17,18). In contrast, RNase R is unmodified in stationary phase cells (17). This post-translational modification leads to tighter binding of tmRNA and SmpB and subsequent proteolytic degradation of exponential phase RNase R, whereas its counterpart in stationary phase binds tmRNA and SmpB weakly and remains stable (17).…”

“…In contrast, RNase R is unmodified in stationary phase cells (17). This post-translational modification leads to tighter binding of tmRNA and SmpB and subsequent proteolytic degradation of exponential phase RNase R, whereas its counterpart in stationary phase binds tmRNA and SmpB weakly and remains stable (17). Acetylation of RNase R is limited to exponential phase because the acetylating activity decreases in late exponential phase and is absent in stationary phase cells (18).…”