title: Clumping factor B, a fibrinogen-binding MSCRAMM (microbial surface components recognizing adhesive matrix molecules) adhesin of Staphylococcus aureus, also binds to the tail region of type I cytokeratin 10.
abstract: The primary habitat of Staphylococcus aureus in humans is the moist squamous epithelium of the anterior nares. We showed previously that S. aureus adheres to desquamated epithelial cells and that clumping factor B (ClfB), a surface-located MSCRAMM (microbial surface components recognizing adhesive matrix molecules) known for its ability to bind to the alpha-chain of fibrinogen, is partly responsible (O'Brien, L. M., Walsh, E. J., Massey, R. C., Peacock, S. J., and Foster, T. J. (2002) Cell. Microbiol. 4, 759-770). We identified cytokeratin 10 (K10) as the ligand recognized by ClfB. Here we have shown that purified recombinant human and murine K10 immobilized on a plastic surface supports adherence of S. aureus in a ClfB-dependent manner. Furthermore, the recombinant A domain of ClfB (rClfB 45-542) bound to immobilized K10 dose-dependently and saturably. Subdomains of human and murine K10 were expressed and purified. The N-terminal head domain (residues 1-145) did not support the binding of rClfB or adherence of S. aureus ClfB+. In contrast, the C-terminal tail domains (human rHK10 452-593, mouse rMK10 454-570) promoted avid binding and adherence. Isothermal titration microcalorimetry and intrinsic tryptophan fluorescence experiments gave dissociation constants for rClfB 45-542 binding to rMK10 454-570 of 1.4 and 1.7 microM, respectively. The tail region of K10 is composed largely of quasi-repeats of Tyr-(Gly/Ser)n. A synthetic peptide corresponding to a typical glycine loop (YGGGSSGGGSSGGY; Y-Y loop peptide) inhibited the adherence of S. aureus ClfB+ to immobilized MK10 to a level of 80%, whereas control peptides had no effect. The KD of rClfB 45-542 for the Y-Y loop peptide was 5.3 microm by intrinsic tryptophan fluorescence. Thus ClfB binds to the glycine loop region of the tail domain of keratin 10 where there are probably multiple binding sites. Binding is discussed in the context of the dock-lock-latch model for MSCRAMM-ligand interactions. We provide an explanation for the molecular basis for S. aureus adherence to the squamous epithelium and suggest that nasal colonization might be prevented by reagents that inhibit this interaction.
authors.affiliation: Department of Microbiology, Moyne Institute of Preventive Medicine, Trinity College Dublin, Dublin 2, Ireland.
authors.forename: Evelyn J
authors.initials: EJ
authors.lastname: Walsh
authors.forename: Louise M
authors.initials: LM
authors.lastname: O'Brien
authors.forename: Xiaowen
authors.initials: X
authors.lastname: Liang
authors.forename: Magnus
authors.initials: M
authors.lastname: Hook
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Foster
mesh_terms: D018829:Adhesins, Bacterial / Q000737:chemistry / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D001422:Bacterial Adhesion; D001665:Binding Sites; D015153:Blotting, Western; D002151:Calorimetry; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D005998:Glycine / Q000737:chemistry; D006801:Humans; D007633:Keratins / Q000737:chemistry*; D008024:Ligands; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence; D013211:Staphylococcus aureus / Q000378:metabolism*; D014364:Tryptophan / Q000737:chemistry
chemical_list: D018829:Adhesins, Bacterial; D017931:DNA Primers; D018076:DNA, Complementary; D008024:Ligands; D010455:Peptides; D011994:Recombinant Proteins; C434058:clfB protein, Staphylococcus aureus; D007633:Keratins; D014364:Tryptophan; D005998:Glycine