Arrangement of the translocator of the autotransporter adhesin involved in diffuse adherence on the bacterial surface.
Autotransporters of gram-negative bacteria are single-peptide secretion systems that consist of a functional N-terminal alpha-domain ("passenger") fused to a C-terminal beta-domain ("translocator"). How passenger proteins are translocated through the outer membrane has not been resolved, and at present essentially three different models are discussed. In the widely accepted "hairpin model" the passenger proteins are translocated through a channel formed by the beta-barrel of the translocator that is integrated in the outer membrane. This model has been challenged by a recent proposal for a general autotransporter model suggesting that there is a hexameric translocation pore that is generated by the oligomerization of six beta-domains. A third model suggests that conserved Omp85 participates in autotransporter integration and passenger protein translocation. To examine these models, in this study we investigated the presence of putative oligomeric structures of the translocator of the autotransporter adhesin involved in diffuse adherence (AIDA) in vivo by cross-linking techniques. Furthermore, the capacity of isolated AIDA fusion proteins to form oligomers was studied in vitro by several complementary analytical techniques, such as analytical gel filtration, electron microscopy, immunogold labeling, and cross-linking of recombinant autotransporter proteins in which different passenger proteins were fused to the AIDA translocator. Our results show that the AIDA translocator is mostly present as a monomer. Only a fraction of the AIDA autotransporter was found to form dimers on the bacterial surface and in solution. Higher-order structures, such as hexamers, were not detected either in vivo or in vitro and can therefore be excluded as functional moieties for the AIDA autotransporter.
Müller
Daniel
D
Institut für Infektiologie, Zentrum für Molekularbiologie der Entzündung (ZMBE), Westfälische Wilhelms-Universität, Von-Esmarch-Str. 56, D-48149 Münster, Germany.
Benz
Inga
I
Tapadar
Damini
D
Buddenborg
Christian
C
Greune
Lilo
L
Schmidt
M Alexander
MA
D018830:Adhesins, Escherichia coli / Q000378:metabolism*; D001422:Bacterial Adhesion; D002352:Carrier Proteins / Q000737:chemistry*; D002850:Chromatography, Gel; D019281:Dimerization; D029968:Escherichia coli Proteins / Q000737:chemistry*; D008854:Microscopy, Electron; D017510:Protein Folding; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000737:chemistry
C075559:AIDA-I protein, E coli; D018830:Adhesins, Escherichia coli; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins