The antibacterial toxin colicin N binds to the inner core of lipopolysaccharide and close to its translocator protein.
Bottom Line: Delipidated LPS (LPS(Δ) (LIPID) ) shows weaker binding; and thus full affinity requires the lipid component.The site of LPS binding means that ColN will preferably bind at the interface and thus position itself close to the surface of its translocon component, OmpF.ColN is, currently, unique among colicins in requiring LPS and, combined with previous data, this implies that the ColN translocon is distinct from those of other known colicins.
Affiliation: Centre for Bacterial Cell Biology, Institute for Cell and Molecular Biosciences, Faculty of Medical Sciences, Newcastle University, Framlington Place, Newcastle-upon-Tyne, NE2 4HH, UK.Show MeSH
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Mentions: SPR was used to identify which regions of ColN were responsible for LPS binding. Various C‐terminally histidine‐tagged ColN domain mutants were injected at equal molar concentrations over a Ni2+ charged NTA surface followed by either Rc LPS (Fig. 3A) or Rd LPS (Fig. 3B). The data demonstrate that all R‐domain containing constructs (i.e. ColN, ColN‐TR and ColN‐R) bind to Rc LPS while ColN‐T and ColN‐P do not (Fig. 3A). LPS dissociates more slowly from ColN or ColN‐R than from ColN‐TR which nevertheless binds significant amounts of LPS. Thus ColN‐R serves as the minimal Rc LPS binding unit and it binds with a similar kinetic profile in both the isolated construct and the full‐length ColN protein. In agreement with the genetic screen (Sharma et al., 2009) none of the constructs tested bound to Rd LPS (Fig. 3B). All the colicin constructs possess a high net positive charge (pI = ∼ 9.0) and show weak, non‐specific, electrostatic binding to any negatively charged LPS which is revealed by small changes in response units (RUs) with Rd LPS (Fig. 3B) and for ColN‐T and ColN‐P with Rc LPS.
Affiliation: Centre for Bacterial Cell Biology, Institute for Cell and Molecular Biosciences, Faculty of Medical Sciences, Newcastle University, Framlington Place, Newcastle-upon-Tyne, NE2 4HH, UK.