Structural determinants of endopilus assembly, stability and functional specificity in bacterial type II secretion

Area of Science:

• Microbiology
• Structural Biology
• Biochemistry

Background:

• Gram-negative bacteria utilize the type II secretion system (T2SS) for extracellular protein transport.
• The T2SS nanomachine assembles a periplasmic polymer called the endopilus, essential for effector translocation.
• Endopili share structural similarities with type IV pili but possess unique features, including a calcium-binding site.

Purpose of the Study:

• To investigate the structural basis of endopilus assembly, stability, and functional specificity.
• To compare the Out T2SS from Dickeya dadantii and the Pul T2SS from Klebsiella oxytoca.
• To identify molecular determinants of secretion specificity and endopilus stability.

Main Methods:

• Comparative analysis of Out and Pul T2SS.
• Nuclear Magnetic Resonance (NMR) structure determination of calcium-bound OutG monomer.
• Cryo-electron microscopy (cryo-EM) structure determination of OutG and PulG endopili.
• Integration of structural, mutational, and biophysical analyses with in vivo assays.

Main Results:

• Determined high-resolution structures of endopili from D. dadantii and K. oxytoca.
• Identified conserved and variable regions within major pilin subunits (OutG and PulG).
• Elucidated molecular determinants governing endopilus stability and secretion specificity.

Conclusions:

• Subtle sequence variations in conserved nanomachines like T2SS contribute to functional optimization.
• Endopilus structure and stability are finely tuned for adaptation to specific ecological niches.
• Understanding T2SS provides insights into bacterial pathogenesis and potential therapeutic targets.