Surface display of the receptor-binding domain of the F17a-G fimbrial adhesin through the autotransporter AIDA-I leads to permeability of bacterial cells

1 Onderzoeksgroep Genetische Virologie, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium 2 Structural Biology Brussels, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium 3 Structural Biology Brussels, Department of Molecular and Cellular Interactions, VIB, B-1050 Brussels, Belgium Correspondence Jean-Pierre Hernalsteens jphernal{at}vub.ac.be 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 P BAD 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 β -galactosidase, a bacterial protein expressed in the cytoplasm, allowed the simultaneous detection of antigen expression and permeability in individual cells. Abbreviations: AIDA-I, adhesin involved in diffuse adherence; AFM, atomic force microscopy; OM, outer membrane Present address: Afdeling Gentechnologie, KU Leuven, Kasteelpark Arenberg 30, B-3001 Heverlee, Belgium.