Clumping Factor B, a Fibrinogen-Binding MSCRAMM adhesin of Staphylococcus aureus, also Binds to the Tail Region of Type I Cytokeratin 10

The primary habitat of Staphylococcus aureus in man 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, a surface located MSCRAMM known for its ability to bind to the α-chain of fibrinogen, is in part 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 also identified cytokeratin 10 (K10) as the ligand recognized by ClfB. In this paper 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 (rClfB45-542) 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 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 rClfB45-542 binding to rMK10 454-570 of 1.4 μM and 1.7 μM, 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 inhibitory effect. The KD of rClfB45-542 for the Y-Y loop peptide was shown to be 5.3 μM by intrinsic trytophan fluorescence. Thus ClfB binds to the glycine loop region of the tail domain of keratin 10 where there are likely to be multiple binding sites. Binding to this region is discussed in the context of the dock-lock-latch model for MSCRAMM-ligand interactions. This paper provides an explanation for the molecular basis for S.aureus adherence to the by gest on N ovem er 7, 2017 hp://w w w .jb.org/ D ow nladed from