Insights into the complex formation of a trimeric autotransporter adhesin with a peptidoglycan-binding periplasmic protein

Abstract

AbstractTrimeric autotransporter adhesin (TAA) is an outer membrane (OM) protein that is widely distributed in gram-negative bacteria and is involved primarily in adhesion to biotic and abiotic surfaces, cell agglutination, and biofilm formation. TAAs are secreted onto the OM by the type Vc secretion system (SS). Because the interactions between TAAs and chaperones or special assistant proteins during secretion are short-lived, it is thought that TAAs reside on the OM without forming complexes with other proteins after secretion. In this study, we aimed to clarify the interactions between anAcinetobacterTAA, AtaA, and a peptidoglycan (PG)-binding periplasmic protein, TpgA, and to identify additional roles of TpgA based on these interactions. Pull-down assays using recombinant proteins identified the interacting domains. X-ray crystallography revealed the A3B3 heterohexameric complex structure of the N-terminal domain of TpgA with the transmembrane domain of AtaA and showed that both electrostatic and hydrophobic interactions contribute to stable complex formation. Bioinformatic analysis suggested that the TAA-TpgA complex is formed in a wide range of species that harbor thetaa-tpgAgene cassette in their genome. Furthermore, the absence of TpgA increased the release of AtaA from the cell surface, suggesting that TpgA prevents the release of TAA from the cell surface via its anchoring to the PG. We propose that the TAAs that form a complex with TpgA be assigned to type Vc2SS, a new subtype of type Vc SS.SignificanceGram-negative bacteria have specialized secretion systems (SSs) that translocate molecules from the cytoplasm to the extracellular space. This is the first report directly demonstrating that the transmembrane domain of a trimeric autotransporter adhesin (TAA), which is secreted by the type Vc SS, forms a stable complex with a peptidoglycan-binding periplasmic protein. This complex prevents the release of TAA from the cell surface. These results suggest that the secretion and architecture of some TAAs are more complex than was previously understood. Our work provides structural and functional insights into bacterial SSs that are important not only in microbiology but also for medical and bioengineering applications.