Loss of outer membrane proteins without inhibition of lipid export in an Escherichia coli YaeT mutant.
Escherichia coli yaeT encodes an essential, conserved outer membrane (OM) protein that is an ortholog of Neisseria meningitidis Omp85. Conflicting data with N. meningitidis indicate that Omp85 functions either in assembly of OM proteins or in export of OM lipids. The role of YaeT in E. coli was investigated with a new temperature-sensitive mutant harboring nine amino acid substitutions. The mutant stops growing after 60 min at 44 degrees C. After 30 min at 44 degrees C, incorporation of [35S]methionine into newly synthesized OM proteins is selectively inhibited. Synthesis and export of OM phospholipids and lipopolysaccharide are not impaired. OM protein levels are low, even at 30 degrees C, and the buoyant density of the OM is correspondingly lower. By Western blotting, we show that levels of the major OM protein OmpA are lower in the mutant in whole cells, membranes, and the growth medium. SecA functions as a multicopy suppressor of the temperature-sensitive phenotype and partially restores OM proteins. Our data are consistent with a critical role for YaeT in OM protein assembly in E. coli.
Doerrler
William T
WT
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
Raetz
Christian R H
CR
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D001692:Biological Transport; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D002855:Chromatography, Thin Layer; D017931:DNA Primers / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000502:physiology; D015723:Gene Library; D005838:Genotype; D050356:Lipid Metabolism; D008070:Lipopolysaccharides / Q000378:metabolism; D008957:Models, Genetic; D009154:Mutation; D009345:Neisseria meningitidis / Q000378:metabolism; D010641:Phenotype; D010743:Phospholipids / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D016133:Polymerase Chain Reaction; D013696:Temperature; D013997:Time Factors
D001425:Bacterial Outer Membrane Proteins; C503914:BamA protein, E coli; D017931:DNA Primers; D029968:Escherichia coli Proteins; D008070:Lipopolysaccharides; D010743:Phospholipids