H pilin cyclisation and pilus biogenesis are promiscuous but electrostatic perturbations impair conjugation efficiency
File(s)
Published version
Author(s)
Type
Journal Article
Abstract
During conjugation, plasmid DNA is transferred from donor to recipient bacteria via the plasmid-encoded mating pilus, formed as thin helical assemblies of polymerised pilin subunits. In the IncHI1 R27 plasmid-encoded pilus, the TrhA pilin undergoes cyclisation (via a peptide bond between Gly1 and Asp69), essential for conjugation. Gly1 and Asp69 are exposed on the pilus surface and conserved in all TrhA pilins in the Plascad database. Substituting Asp69 with Asn, Ala, Gly, or Arg does not prevent cyclisation or pilus formation, which remains structurally indistinguishable from the wild type. Conjugation efficiency of the Asp69 substitutions across multiple recipient species correlates with side chain size, in the order Asp69Asn > Asp69Ala > Asp69Gly. However, Asp69Arg, as well as Asp69Lys and Gly1Lys substitutions abolish conjugation, likely due to the positively charged pilus surface (opposite to the wild-type negative charge) forming unfavourable electrostatic interactions with the recipient outer membrane’s inner leaflet, composed solely of zwitterionic phosphatidylethanolamine (PE). Consistently, conjugation is rescued in recipients lacking PE. These findings indicate strong selective pressure to maintain Gly1 and Asp69, as efficient DNA transfer depends on precise electrostatic and steric constraints of the pilus surface.
Date Issued
2026-03-27
Date Acceptance
2026-02-02
Citation
Nature Communications, 2026, 17
ISSN
2041-1723
Publisher
Nature Portfolio
Journal / Book Title
Nature Communications
Volume
17
Copyright Statement
© The Author(s) 2026. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.
License URL
Identifier
10.1038/s41467-026-69599-3
Publication Status
Published
Article Number
ARTN 2888
Date Publish Online
2026-02-18