Surface display of the receptor-binding domain of the F17a-G fimbrial adhesin through the autotransporter AIDA-I leads to permeability of bacterial cells.
Surface exposure of antigens on bacterial cells can be critical for eliciting an effective antibody response. Therefore, we investigated the cellular localization of the fimbrial F17a-G receptor-binding domain, fused to the translocator domain of the AIDA-I autotransporter. Synthesis of the fusion protein, under the control of the L-arabinose-inducible PBAD promoter, was shown to permeabilize Escherichia coli K-12 and Salmonella enterica serovar Typhimurium cells. The presence of permeable cells interfered with several methods that are typically used to determine surface exposure of proteins, such as protease treatment and whole-cell ELISA. Double immunofluorescence microscopy, using a second antibody directed against beta-galactosidase, a bacterial protein expressed in the cytoplasm, allowed the simultaneous detection of antigen expression and permeability in individual cells.
Van Gerven
Nani
N
Onderzoeksgroep Genetische Virologie, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium.
Sleutel
Mike
M
Structural Biology Brussels, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium.
Deboeck
Francine
F
Onderzoeksgroep Genetische Virologie, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium.
De Greve
Henri
H
Structural Biology Brussels, Department of Molecular and Cellular Interactions, VIB, B-1050 Brussels, Belgium.|Structural Biology Brussels, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium.
Hernalsteens
Jean-Pierre
JP
Onderzoeksgroep Genetische Virologie, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium.
D018829:Adhesins, Bacterial / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018830:Adhesins, Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002463:Cell Membrane Permeability; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D050296:Microbial Viability; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012486:Salmonella typhimurium / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
C075559:AIDA-I protein, E coli; D018829:Adhesins, Bacterial; D018830:Adhesins, Escherichia coli; D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; C478166:F17-G protein, E coli; D011993:Recombinant Fusion Proteins