9N32

PilU from Pseudomonas aeruginosa: C3ocococ conformation

Summary for 9N32

• Entry DOI: 10.2210/pdb9n32/pdb
• Descriptor: PilU protein (1 entity in total)
• Functional Keywords: pilu, pilt, type iv pilus, t4p, t4ap, pseudomonas, aeruginosa, motility, atpase, pilus, motor protein
• Biological source: Pseudomonas aeruginosa
• Total number of polymer chains: 4
• Total formula weight: 179040.47

Authors

McCallum, M.,Howell, P.L.,Burrows, L. (deposition date: 2025-01-29, release date: 2025-12-10)

Primary citation

Barnshaw, R.,Harvey, H.,McCallum, M.,Lazarou, T.,Nguyen, S.,Qaderi, I.,Tran, V.,Roberge, N.,Geiger, C.,O'Toole, G.A.,Howell, P.L.,Burrows, L.L.
Separation of Pseudomonas aeruginosa type IV pilus-dependent twitching motility and surface-sensing responses.
Mbio, 16:e0252125-e0252125, 2025

PubMed Abstract: Many functions of type IVa pili, including twitching motility and surface sensing, depend on dynamic cycles of filament assembly and disassembly powered by the cytoplasmic ATPases PilB, PilT, and PilU. Deletion of results in loss of twitching and pilus-specific bacteriophage susceptibility, while non-twitching mutants remain susceptible to pilus-specific phages, indicating that they still produce retractable pili. mutants have high basal levels of the secondary messenger cyclic AMP (cAMP) that normally increases following surface contact, suggesting aberrant surface sensing. To better understand PilU's role in pilus biology, we solved its X-ray crystal structure and used phylogenetic analyses to identify conserved differences between PilT and PilU. Chemical mutagenesis followed by whole-genome sequencing was used to identify suppressors in the Δ mutant background that restored twitching motility. The mutations mapped to the major pilin, PilA, or the pilus tip adhesin, PilY1. Both the position and nature of the substitutions in PilA impacted restoration of motility, and it was dependent on functional PilT. Complementation of most suppressors with PilU further increased motility, while the expression of wild-type PilA decreased motility in a dose-dependent manner. Notably, cAMP levels remained elevated in most twitching suppressor mutants, showing that surface sensing and motility can be uncoupled. Together, our data suggest that the bacterial response to surfaces reflects a complex interaction of PilU function with specific alleles of PilY1 and PilA that together modulate pilus dynamics and function.

PubMed: 41055394
DOI: 10.1128/mbio.02521-25

Experimental method

X-RAY DIFFRACTION (4.54 Å)