Functional characterization of Escherichia coli GlpG and additional rhomboid proteins using an aarA mutant of Providencia stuartii.
The Providencia stuartii AarA protein is a member of the rhomboid family of intramembrane serine proteases and required for the production of an extracellular signaling molecule that regulates cellular functions including peptidoglycan acetylation, methionine transport, and cysteine biosynthesis. Additional aarA-dependent phenotypes include (i) loss of an extracellular yellow pigment, (ii) inability to grow on MacConkey agar, and (iii) abnormal cell division. Since these phenotypes are easily assayed, the P. stuartii aarA mutant serves as a useful host system to investigate rhomboid function. The Escherichia coli GlpG protein was shown to be functionally similar to AarA and rescued the above aarA-dependent phenotypes in P. stuartii. GlpG proteins containing single alanine substitutions at the highly conserved catalytic triad of asparagine (N154A), serine (S201A), or histidine (H254A) residues were nonfunctional. The P. stuartii aarA mutant was also used as a biosensor to demonstrate that proteins from a variety of diverse sources exhibited rhomboid activity. In an effort to further investigate the role of a rhomboid protein in cell physiology, a glpG mutant of E. coli was constructed. In phenotype microarray experiments, the glpG mutant exhibited a slight increase in resistance to the beta-lactam antibiotic cefotaxime.
Clemmer
Katy M
KM
Laboratories of Microbial Pathogenesis, Atlanta VA Medical Center, Atlanta, Georgia, USA.
Sturgill
Gwen M
GM
Veenstra
Alexander
A
Rather
Philip N
PN
D000362:Agar; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics*; D015374:Biosensing Techniques / Q000379:methods*; D002439:Cefotaxime / Q000494:pharmacology; D002455:Cell Division; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000502:physiology; D004351:Drug Resistance; D010450:Endopeptidases / Q000235:genetics* / Q000502:physiology; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000502:physiology; D008565:Membrane Proteins / Q000235:genetics* / Q000502:physiology; D009154:Mutation; D010860:Pigments, Biological / Q000096:biosynthesis; D011532:Providencia / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D011995:Recombination, Genetic; D012097:Repressor Proteins / Q000235:genetics*; D012697:Serine Endopeptidases / Q000378:metabolism
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C102898:GlpG protein, E coli; D008565:Membrane Proteins; D010860:Pigments, Biological; D012097:Repressor Proteins; C088713:aarA protein, Providencia stuartii; D000362:Agar; D010450:Endopeptidases; D012697:Serine Endopeptidases; D002439:Cefotaxime