Abstract
Although it is normally an innocuous part of the human skin microbiota, Staphylococcus epidermidis has emerged as a major nosocomial pathogen, and implanted foreign materials are an essential risk factor for the development of an infection. The extraordinary efficiency of S. epidermidis to colonize artificial surfaces is particularly related to the ability to form biofilms. Biofilm formation itself critically depends on stable pathogen binding to extracellular host matrix components, e.g. fibronectin (Fn), covering inserted devices in vast amounts. Extracellular matrix binding protein (Embp) and its subdomains referred to as the F-repeat and the FG-repeat are critical for adherence of S. epidermidis to surface-immobilized Fn. Embp-Fn interactions preferentially occur with surface-bound, but not folded, globular Fn via binding to the F3 domain. High-resolution structure analysis of F- and FG-repeats revealed that both repeats are composed of two tightly connected triple α-helix bundles, exhibiting an elongated but rather rigid structural organization in solution. Both F- and FG-repeat possess Fn-binding capacity via interactions with type III subdomain FN12, involving residues within the C and F β-sheet. FN12 essentially supports stability of the globular Fn state, and thus these findings reasonably explain why Embp-mediated interaction of S. epidermidis necessitates Fn surface immobilization. Thus, Embp employs an uncharacterized bacterial Fn-binding mechanism to promote staphylococcal adherence.IMPORTANCEStaphylococcus epidermidis is a leading pathogen in implant-associated hospital infections. The pathogenesis critically depends on bacterial binding to ECM components, specifically fibronectin (Fn). The cell surface-localized, 1-MDa extracellular matrix binding protein (Embp) is essentially characterized by 10 F- and 40 FG-repeats. These repetitive units, each characterized by two α-helical bundles, organize themselves in a rigid, elongated form. Embp binds preferentially to surface-localized but not soluble Fn, with both F- and FG-repeats being sufficient for Fn binding and resulting bacterial adherence. Binding preferentially involves Fn type III domain, specifically residues of FN12 β-sheets C and F. Both play key role in stabilizing the globular Fn conformation, explaining the necessity of Fn surface immobilization for a subsequent interaction with Embp. In comparison to many other bacterial Fn-binding proteins using the Fn N terminus, Embp employs a previously undescribed mechanism supporting the adhesion of S. epidermidis to surface-immobilized Fn.
Highlights
It is normally an innocuous part of the human skin microbiota, Staphylococcus epidermidis has emerged as a major nosocomial pathogen, and implanted foreign materials are an essential risk factor for the development of an infection
In order to test the effect of extracellular matrix binding protein (Embp) production on cell surface recruitment of soluble Fn, S. epidermidis 1585Pxyl/tetembp, a strain in which the embp expression can be switched on and off via a tetracycline inducible promoter [22], and the embp knockout mutant S. epidermidis 1585Δembp was suspended in phosphate-buffered saline (PBS) containing Fn (10 g/ml) for 1 h
Interactions with host extracellular matrix (ECM) components are a crucial step in colonization and infection establishment of many bacteria, and well over 100 bacterial cell surface proteins with Fn-binding activity have been identified so far [34]
Summary
It is normally an innocuous part of the human skin microbiota, Staphylococcus epidermidis has emerged as a major nosocomial pathogen, and implanted foreign materials are an essential risk factor for the development of an infection. Extracellular matrix binding protein (Embp) and its subdomains referred to as the F-repeat and the FGrepeat are critical for adherence of S. epidermidis to surface-immobilized Fn. Embp-Fn interactions preferentially occur with surface-bound, but not folded, globular Fn via binding to the F3 domain. The cell surface-localized, 1-MDa extracellular matrix binding protein (Embp) is essentially characterized by 10 F- and 40 FG-repeats These repetitive units, each characterized by two ␣-helical bundles, organize themselves in a rigid, elongated form. Molecular work from the past decades revealed that S. epidermidis biofilm formation is a multistep process involving a plethora of bacterial molecules, all integrated into complex regulatory circuits [7, 8] Many of these factors are well-characterized adhesins (e.g., polysaccharide intercellular adhesin [PIA], accumulation associated protein [Aap], extracellular DNA [eDNA] [9]) fostering intercellular adhesion, cell aggregation and, the establishment of a multicellular biofilm architecture [10]. Crystal structure analysis of core Embp units and their functional characterization provide novel insights into a unique bacterial mechanism contributing to Fn binding and surface colonization
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