Lookup NU author(s): Jonathan Pointon,
Dr Emily Abbot,
Professor Barry Hirst,
Professor Janet Wilson,
Dr Mark Banfield,
Professor Michael Kehoe
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Adhesive pili on the surface of the M1 Streptococcus pyogenes strain SF370 are composed of a major 'backbone' subunit (Spy0128) and two minor subunits (Spy0125 and Spy0130), joined covalently by a pilin-polymerase (Spy0129). Previous studies using recombinant proteins showed that both minor subunits bind to human pharyngeal (Detroit) cells (Manetti, A.G., et al, 2007, Mol Microbiol 64:968-83), suggesting both may act as pilus-presented adhesins. While confirming these binding properties, studies described here indicate that Spy0125 is the pilus-presented adhesin and that Spy0130 has a distinct role as a 'wall-linker'. Pili were localised predominantly to cell-wall fractions of the wild-type S. pyogenes parent strain and a spy0125 deletion mutant. In contrast, they were found almost exclusively in culture supernatants in both spy0130 and srtA deletion mutants, indicating that the housekeeping sortase (SrtA) attaches pili to the cell-wall using Spy0130 as a linker protein. Adhesion assays with antisera specific for individual subunits showed that only anti-rSpy0125 sera inhibited adhesion of wild-type S. pyogenes to human keratinocytes and tonsil epithelium to a significant extent. Spy0125 was localised to the tip of pili by a combination of mutant analysis and LC-MS/MS analysis of purified pili. Assays comparing parent and mutant strains confirmed its role as the adhesin. Unexpectedly, apparent spontaneous cleavage of a labile, proline-rich (8/14 residues), sequence separating the N-terminal approximately 1/3 and C-terminal approximately 2/3 of Spy0125 leads to loss of the N-terminal region, but analysis of internal spy0125 deletion mutants confirmed that this has no significant effect on adhesion.
Author(s): Smith WD, Pointon JA, Abbot E, Kang HJ, Baker EN, Hirst BH, Wilson JA, Banfield MJ, Kehoe MA
Publication type: Article
Journal: Journal of Bacteriology
Print publication date: 19/07/2010
ISSN (print): 0021-9193
ISSN (electronic): 1098-5530
Publisher: American Society for Microbiology
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