The binding of Pseudomonas aeruginosa pili to glycosphingolipids is a tip‐associated event involving the C‐terminal region of the structural pilin subunit

Abstract

Pili are one of the adhesins of Pseudomonas aeruginosa that mediate adherence to epithelial cell-surface receptors. The pili of P. aeruginosa strains PAK and PAO were examined and found to bind gangliotetraosyl ceramide (asialo-GM1) and, to a lesser extend, II3N-acetylneuraminosylgangliotetraosyl ceramide (GM1) in solid-phase binding assays. Asialo-GM1, but not GM1, inhibited both PAK and PAO pili binding to immobilized asialo-GM1 on the microtitre plate. PAO pili competitively inhibited PAK pili binding to asialo-GM1, suggesting the presence of a structurally similar receptor-binding domain in both pilus types. The interaction between asialo-GM1 and pili occurs at the pilus tip as asialo-GM1 coated colloidal gold only decorates the tip of purified pili. Three sets of evidence suggest that the C-terminal disulphide-bonded region of the Pseudomonas pilin is exposed at the tip of the pilus: (i) immunocytochemical studies indicate that P. aeruginosa pili have a basal-tip structural differentiation where the monoclonal antibody (mAb) PK3B recognizes an antigenic epitope displayed only on the basal ends of pili (produced by shearing) while the mAb PK99H, whose antigenic epitope resides in residues 134-140 (Wong et al., 1992), binds only to the tip of PAK pili; (ii) synthetic peptides, PAK(128-144)ox-OH and PAO(128-144)ox-OH, which correspond to the C-terminal disulphide-bonded region of Pseudomonas pilin are able to bind to asialo-GM1 and inhibit the binding of pili to the glycolipid; (iii) PK99H was shown to block PAK pilus binding to asialo-GM1.(ABSTRACT TRUNCATED AT 250 WORDS)

Mutational analysis of receptor binding mediated by the Dr family of Escherichia coli adhesins

“…Synthetic peptides have been used to define receptorbinding domains of several adhesins (Leininger et al, 1991;Lee et al, 1994;Patti et al, 1995). In our study, we tested the effects on DAF binding of three synthetic peptides corresponding to the N-terminal region of DraE and observed an inability of these peptides to inhibit binding, despite the fact that they covered a region involved in the binding process according to the data obtained with the point mutants.…”

Laboratory and clinical Pseudomonas aeruginosa strains do not bind glycosphingolipids in vitro or during type IV pili-mediated initial host cell attachment

“…GSL recognition is a function of a C-terminal disulfide-linked domain at the pilus tip (Lee et al, 1994). We see that the T4P tips are clearly visible on eukaryotic-cell-attached P. aeruginosa, which should therefore be available for GSL binding.…”

“…This is not merely aggregation but rather a 'T4P tip to T4P tip' interaction to give a network of attached bacteria. The pilus tip is the Gg 3 /Gg 4 binding domain (Lee et al, 1994), and thus interpilus binding may prevent bacterial Gg 3 /Gg 4 binding. A further possibility is the disassembly of the pilus subunits once detached from the bacterium, and the augmentary or differential binding of the smaller fragments, each of which contains a receptorbinding-tip domain.…”

“…The finding that P. aeruginosa expressing type IV pili belonging to group 1 are significantly overrepresented among isolates from the CF population (Kus et al, 2004) prompted the current study to determine whether differential receptor binding specificity might explain this epidemiological correlation. The binding of T4P in in vitro assays to the glycosphingolipids (GSLs) gangliotriaosylceramide (GalNAcb1-4Galb-4Glc ceramide, Gg 3 ) and gangliotetraosylceramide (Galb1-3GalNAcb1-4Gal b-4Glc ceramide, Gg 4 ) (also commonly termed asialoGM2 and asialoGM1, respectively) (Comolli et al, 1999b;Gupta et al, 1994;Lee et al, 1994;Saiman & Prince, 1993;Schweizer et al, 1998) has implicated these molecules as cellular receptors for adhesion of P. aeruginosa (Baker et al, 1990;Hazlett et al, 1993;Krivan et al, 1988a;Ramphal et al, 1991a). These GSLs contain the common sequence GalNAcb1-4Gal, which has been defined as the minimal T4P recognition epitope or adhesintope (Campbell et al, 1997;Lee et al, 1996).…”

Neisseria gonorrhoeae PilV, a type IV pilus-associated protein essential to human epithelial cell adherence

“…In contrast, Tfp-mediated adherence exhibited by Pseudomonas aeruginosa has been proposed to involve a receptor-binding site within the C terminus of the pilin subunit (9). Tfp also enhance colonization of human epithelium by enteropathogenic strains of Escherichia coli (10) and Vibrio cholerae (11), but is unclear whether specific adhesin-receptor interactions are involved in these species.…”