Structure-guided disruption of the pseudopilus tip complex inhibits the Type II secretion in Pseudomonas aeruginosa

Area of Science:

• Microbiology
• Structural Biology
• Molecular Biology

Background:

• The Type II secretion system (T2SS) in Pseudomonas aeruginosa is vital for translocating protein virulence factors essential for infection.
• The T2SS relies on pseudopilins assembling into a pseudopilus, acting as a piston to secrete exoproteins.

Purpose of the Study:

• To determine the structure and function of pseudopilin complexes involved in the T2SS.
• To investigate the role of minor pseudopilins XcpV and XcpW in pseudopilus assembly and function.
• To develop novel inhibitory peptides targeting the T2SS based on structural insights.

Main Methods:

• Structure determination of XcpVWX and XcpVW complexes.
• Functional analysis of pseudopilin mutants.
• Small-angle X-ray scattering (SAXS) for pseudopilus tip architecture.
• In vitro and in vivo inhibition assays for developed peptides.
• In vivo virulence assays using Caenorhabditis elegans.

Main Results:

• Two minor pseudopilins, XcpV and XcpW, form an indispensable core complex at the pseudopilus tip.
• The absence of XcpV or XcpW renders the T2SS non-functional.
• SAXS experiments elucidated the pseudopilus tip architecture and established a working model.
• Structure-based inhibitory peptides disrupted the XcpVW complex and pseudopilus tip in vitro.
• These peptides inhibited T2SS function in vivo and significantly reduced P. aeruginosa virulence in C. elegans.

Conclusions:

• The XcpVW core complex is essential for T2SS functionality in P. aeruginosa.
• Structure-based peptide inhibitors offer a promising strategy for combating P. aeruginosa infections by targeting the T2SS.
• This study provides a mechanistic understanding of pseudopilus assembly and opens avenues for therapeutic interventions.