Channel formation by FhaC, the outer membrane protein involved in the secretion of the Bordetella pertussis filamentous hemagglutinin.
Many virulence factors of pathogenic microorganisms are presented at the cell surface. However, protein secretion across the outer membrane of Gram-negative bacteria remains poorly understood. Here we used the extremely efficient secretion of the Bordetella pertussis filamentous hemagglutinin (FHA) to decipher this process. FHA secretion requires a single specific accessory protein, FhaC, the prototype of a family of proteins necessary for the extracellular localization of various virulence proteins in Gram-negative bacteria. We show that FhaC is heat-modifiable and localized in the outer membrane. Circular dichroism spectra indicated that FhaC is rich in beta-strands, in agreement with structural predictions for this protein. We further demonstrated that FhaC forms pores in artificial membranes, as evidenced by single-channel conductance measurements through planar lipid bilayers, as well as by liposome swelling assays and patch-clamp experiments using proteoliposomes. Single-channel conductance appeared to fluctuate very fast, suggesting that the FhaC channels frequently assume a closed conformation. We thus propose that FhaC forms a specific beta-barrel channel in the outer membrane for the outward translocation of FHA.
Jacob-Dubuisson
F
F
INSERM U447, Institut de Biologie de Lille, Institut Pasteur de Lille, 1 rue Calmette, 59019 Lille Cedex, France. francoise.jacob@pasteur-lille.fr
El-Hamel
C
C
Saint
N
N
Guédin
S
S
Willery
E
E
Molle
G
G
Locht
C
C
D001425:Bacterial Outer Membrane Proteins / Q000302:isolation & purification / Q000502:physiology*; D001483:Base Sequence; D001886:Bordetella pertussis / Q000378:metabolism* / Q000472:pathogenicity; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D017931:DNA Primers; D006388:Hemagglutinins / Q000378:metabolism*; D006358:Hot Temperature; D013347:Subcellular Fractions / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D017931:DNA Primers; D006388:Hemagglutinins; C098052:fhaC protein, Bordetella pertussis