The Translocation Domain in Trimeric Autotransporter Adhesins Is Necessary and Sufficient for Trimerization and Autotransportation

Abstract

Trimeric autotransporter adhesins (TAAs) comprise one of the secretion pathways of the type V secretion system. The mechanism of their translocation across the outer membrane remains unclear, but it most probably occurs by the formation of a hairpin inside the ␤-barrel translocation unit, leading to transportation of the passenger domain from the C terminus to the N terminus through the lumen of the ␤-barrel. We further investigated the phenomenon of autotransportation and the rules that govern it. We showed by coexpressing different Escherichia coli immunoglobulin-binding (Eib) proteins that highly similar TAAs could form stochastically mixed structures (heterotrimers). We further investigated this phenomenon by coexpressing two more distantly related TAAs, EibA and YadA. These, however, did not form heterotrimers; indeed, coexpression was lethal to the cells, leading to elimination of one or another of the genes. However, substituting in either protein the barrel of the other one so that the barrels were identical led to formation of heterotrimers as for Eibs. Our work shows that trimerization of the ␤-barrel, but not the passenger domain, is necessary and sufficient for TAA secretion while the passenger domain is not.

Heterologous Expression of Bartonella Adhesin A in Escherichia coli by Exchange of Trimeric Autotransporter Adhesin Domains Results in Enhanced Adhesion Properties and a Pathogenic Phenotype

“…2B). These results confirm earlier observations showing that an exchange of membrane anchor domains of YadA, Hia, Eib, or UspA1 still allowed surface expression of the respective TAAs (20,38). However, these reports were all focused on TAAs from gammaproteobacteria while the results reported here demonstrate that a domain exchange between TAAs from bacteria belonging to different bacterial subdivisions still enables surface expression of chimeric TAAs.…”

Insights into the autotransport process of a trimeric autotransporter, Yersinia Adhesin A (YadA)

“…A) that is essential for passenger translocation and for trimerization of the β‐barrel of TAAs (Roggenkamp et al , ; Meng et al , ). The linker along with the β‐barrel is sufficient for membrane insertion (Wollmann et al , ; Mikula et al , ), indicating that the linker of TAAs plays an important role in their biogenesis. Molecular dynamics simulations of the translocator domain of the TAA Hia from Haemophilus influenzae has shown that the linker forms stabilizing interactions with the β‐sheet of the barrel, which in turn stabilize the trimeric barrel (Holdbrook et al , ).…”

The Haemophilus ducreyi trimeric autotransporter adhesin DsrA protects against an experimental infection in the swine model of chancroid

“…1B) expressed from a plasmid in E. coli was purified using nickel affinity chromatography. Purity of rNT-DsrA I and its ability to form multimers was assessed by SDS-PAGE and Western blot, as trimer formation is important for structure and function in other TAAs [32-34]. The three preparations of rNT-DsrA I used for the vaccine trials were more than 95% pure by Coomassie staining, save a faint band around 45 kDa, assumed to be the dimer form of rNT-DsrA I (Fig.…”