C-terminal DNA binding stimulates N-terminal phosphorylation of the outer membrane protein regulator OmpR from Escherichia coli.
Expression of the porin genes of Escherichia coli is regulated in part by the osmolarity of the growth medium. The process is controlled by the histidine kinase EnvZ and the response regulator OmpR. We have previously shown that phosphorylation of OmpR increases its affinity for the upstream regulatory regions of ompF and ompC. We now report that, in the presence of DNA, there is a dramatic stimulation in the level of phospho-OmpR. This effect is independent of the source of phosphorylation, i.e., stimulation of phosphorylation is observed with a small phosphorylating agent such as acetyl phosphate or with protein-catalyzed phosphorylation by the kinase EnvZ. The dephosphorylation rate of phospho-OmpR is affected only slightly by the presence of DNA; thus, the increased level is largely caused by an increased rate of phosphorylation. Stimulation of phosphorylation requires specific binding of DNA by OmpR. Occupancy of the DNA binding domain exposes a trypsin cleavage site in the linker, which connects the phosphorylation domain with the DNA binding domain. Our results indicate that when DNA binds in the C terminus, it enhances phosphorylation in the N terminus, and the linker undergoes a conformational change. A generalized mechanism involving a four-state model for response regulators is proposed.
Ames
S K
SK
Department of Molecular Microbiology, L-220 Oregon Health Sciences University, 3181 Southwest Sam Jackson Park Road, Portland, OR 97201-3098, USA.
Frankema
N
N
Kenney
L J
LJ
D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001426:Bacterial Proteins; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins; D018257:Helix-Loop-Helix Motifs; D007700:Kinetics; D008956:Models, Chemical; D009097:Multienzyme Complexes; D010755:Organophosphates / Q000378:metabolism; D010766:Phosphorylation; D018272:Porins / Q000378:metabolism; D011487:Protein Conformation; D013997:Time Factors; D015534:Trans-Activators / Q000737:chemistry* / Q000378:metabolism*; D015533:Transcriptional Activation; D014357:Trypsin / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D009097:Multienzyme Complexes; D010755:Organophosphates; D018272:Porins; D015534:Trans-Activators; C112177:osmolarity response regulator proteins; C011632:acetyl phosphate; D004247:DNA; C057843:envZ protein, E coli; D014357:Trypsin