Secondary structure and 1H, 15 N & 13C resonance assignments of the periplasmic domain of OutG, major pseudopilin from Dickeya dadantii type II secretion system.

The ability to interact and adapt to the surrounding environment is vital for bacteria that colonise various niches and organisms. One strategy developed by Gram-negative bacteria is to secrete exoprotein substrates via the type II secretion system (T2SS). The T2SS is a proteinaceous complex spanning the bacterial envelope that translocates folded proteins such as toxins and enzymes from the periplasm to the extracellular milieu. In the T2SS, a cytoplasmic ATPase elongates in the periplasm the pseudopilus, a non-covalent polymer composed of protein subunits named pseudopilins, and anchored in the inner membrane by a transmembrane helix. The pseudopilus polymerisation is coupled to the secretion of substrates. The T2SS of Dickeya dadantii secretes more than 15 substrates, essentially plant cell wall degrading enzymes. In D. dadantii, the major pseudopilin or the major subunit of the pseudopilus is called OutG. To better understand the mechanism of secretion of these numerous substrates via the pseudopilus, we have been studying the structure of OutG by NMR. Here, as the first part of this study, we report the 1H, 15N and 13C backbone and sidechain chemical shift assignment of the periplasmic domain of OutG and its NMR derived secondary structure.

The gp27-like Hub of VgrG Serves as Adaptor to Promote Hcp Tube Assembly.

Contractile injection systems are multiprotein complexes that use a spring-like mechanism to deliver effectors into target cells. In addition to using a conserved mechanism, these complexes share a common core known as the tail. The tail comprises an inner tube tipped by a spike, wrapped by a contractile sheath, and assembled onto a baseplate. Here, using the type VI secretion system (T6SS) as a model of contractile injection systems, we provide molecular details on the interaction between the inner tube and the spike. Reconstitution into the Escherichia coli heterologous host in the absence of other T6SS components and in vitro experiments demonstrated that the Hcp tube component and the VgrG spike interact directly. VgrG deletion studies coupled to functional assays showed that the N-terminal domain of VgrG is sufficient to interact with Hcp, to initiate proper Hcp tube polymerization, and to promote sheath dynamics and Hcp release. The interaction interface between Hcp and VgrG was then mapped using docking simulations, mutagenesis, and cysteine-mediated cross-links. Based on these results, we propose a model in which the VgrG base serves as adaptor to recruit the first Hcp hexamer and initiates inner tube polymerization.