Date Published: October 22, 2018
Publisher: Public Library of Science
Author(s): Yichen Zhang, Frédérick Faucher, Wenwen Zhang, Shu Wang, Nolan Neville, Keith Poole, Jun Zheng, Zongchao Jia, Matthew R. Parsek.
Pseudomonas aeruginosa utilizes the Type II secretion system (T2SS) to translocate a wide range of large, structured protein virulence factors through the periplasm to the extracellular environment for infection. In the T2SS, five pseudopilins assemble into the pseudopilus that acts as a piston to extrude exoproteins out of cells. Through structure determination of the pseudopilin complexes of XcpVWX and XcpVW and function analysis, we have confirmed that two minor pseudopilins, XcpV and XcpW, constitute a core complex indispensable to the pseudopilus tip. The absence of either XcpV or -W resulted in the non-functional T2SS. Our small-angle X-ray scattering experiment for the first time revealed the architecture of the entire pseudopilus tip and established the working model. Based on the interaction interface of complexes, we have developed inhibitory peptides. The structure-based peptides not only disrupted of the XcpVW core complex and the entire pseudopilus tip in vitro but also inhibited the T2SS in vivo. More importantly, these peptides effectively reduced the virulence of P. aeruginosa towards Caenorhabditis elegans.
Pseudomonas aeruginosa is a Gram-negative bacterium known for its severe pathogenicity. Therapeutic treatment of P. aeruginosa is difficult due to its low outer member permeability, enzymatic modification and efflux of antibiotics. Its ability to catabolize a wide range of organic molecules guarantees its high adaptability and prevalence in ubiquitous environments[2, 3], which makes it one of the most predominant nosocomial pathogens. As an opportunistic pathogen, it tends to preferentially colonize in immunocompromised patients like those suffering from cystic fibrosis, cancer, or AIDS, and cause pneumonia, urinary tract infection and gastrointestinal infection[5–8]. As a result, the World Health Organization has recently deemed P. aeruginosa a Priority 1 pathogen that is in urgent need of new antibiotics.
The Type II secretion system is widely used in Gram-negative bacteria, including various pathogenic species, to secrete a vast range of virulence factors for infection. However, the molecular mechanism by which the T2SS secretes large, folded toxins has not been fully unveiled. Through the structural and functional studies, we have confirmed that the two minor pseudopilins, XcpV and -W, form a core binary complex that bridges the other minor pseudopilins to assemble the pseudopilus tip complex, which is essential for toxin secretion by the T2SS. Our findings in P. aeruginosa coincide with the results discovered in other species, i.e. K. oxytoca, that the pseudopilus tip is essential for the Type II secretory pathway. By showing that the lack of either XcpV or -W in the XcpVW complex resulted in the non-functional T2SS, we have provided experimental evidence that XcpVW binary complex plays an irreplaceable role in the T2SS of P. aeruginosa, which confirms the speculation that XcpVW functions as the core structure in the system[17, 18, 24]. Therefore, the destabilization of the core complex can lead to the inhibition of the T2SS.