The structure of multiple polypeptide domains determines the signal recognition particle targeting requirement of Escherichia coli inner membrane proteins.
The signal recognition particle (SRP) targeting pathway is required for the efficient insertion of many polytopic inner membrane proteins (IMPs) into the Escherichia coli inner membrane, but in the absence of SRP protein export proceeds normally. To define the properties of IMPs that impose SRP dependence, we analyzed the targeting requirements of bitopic IMPs that are structurally intermediate between exported proteins and polytopic IMPs. We found that disruption of the SRP pathway inhibited the insertion of only a subset of bitopic IMPs. Studies on a model bitopic AcrB-alkaline phosphatase fusion protein (AcrB 265-AP) showed that the SRP requirement for efficient insertion correlated with the presence of a large periplasmic domain (P1). As previously reported, perturbation of the SRP pathway also affected the insertion of a polytopic AcrB-AP fusion. Even exhaustive SRP depletion, however, failed to block the insertion of any AcrB derivative by more than 50%. Taken together, these data suggest that many proteins that are normally targeted by SRP can utilize alternative targeting pathways and that the structure of both hydrophilic and membrane-spanning domains determines the degree to which the biogenesis of a protein is SRP dependent.
Newitt
J A
JA
Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Ulbrandt
N D
ND
Bernstein
H D
HD
D000469:Alkaline Phosphatase / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D001483:Base Sequence; D002462:Cell Membrane / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000502:physiology*; D029968:Escherichia coli Proteins; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017384:Sequence Deletion; D018271:Signal Recognition Particle / Q000378:metabolism*
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D018271:Signal Recognition Particle; C043426:inner membrane protein, E coli; D000469:Alkaline Phosphatase