Display Settings:

Format

Send to:

Choose Destination
See comment in PubMed Commons below
J Bacteriol. 2009 Jul;191(13):4365-71. doi: 10.1128/JB.00204-09. Epub 2009 May 1.

The C-terminal domain of AcrA is essential for the assembly and function of the multidrug efflux pump AcrAB-TolC.

Author information

  • 1Department of Chemistry and Biochemistry, University of Oklahoma, Norman, Oklahoma 73019, USA.

Abstract

Periplasmic membrane fusion proteins (MFPs) are essential components of multidrug efflux pumps and type I protein secretion systems of gram-negative bacteria. Located in the periplasm, MFPs function by creating a physical link between inner membrane transporters and outer membrane channels. The most conserved sequence of MFPs is located in their distal C-terminal domain. However, neither the structure nor the function of this domain is known. In this study, we investigated the structural and functional role of the C-terminal domain of Escherichia coli AcrA, a periplasmic component of the multidrug efflux pump AcrAB-TolC. Using trypsin proteolysis, we identified the proteolytically labile sites in the C-terminal domain (amino acid residues 315 to 397) of AcrA in vitro. We next used these sites as a map to evaluate the structural integrity of this domain of AcrA inside the periplasm. We found that the C-terminal domain of AcrA is protected from trypsin when the tripartite efflux pump AcrAB-TolC is assembled. In contrast, this domain remains proteolytically labile in cells producing only one of the AcrB or TolC components of the complex. Site-directed mutagenesis of 12 highly conserved amino acid residues of the C-terminal domain of AcrA showed that a single G363C substitution dramatically impairs the multidrug efflux activity of AcrAB-TolC. The G363C mutant interacts with both AcrB and TolC but fails to properly assemble into a functional complex. We conclude that the C-terminal domain of AcrA plays an important role in the assembly and function of AcrAB-TolC efflux pump.

PMID:
19411330
[PubMed - indexed for MEDLINE]
PMCID:
PMC2698478
Free PMC Article

Images from this publication.See all images (5)Free text

FIG. 1.
FIG. 2.
FIG. 3.
FIG. 4.
FIG. 5.
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Icon for HighWire Icon for PubMed Central
    Loading ...
    Write to the Help Desk