The giant staphylococcal protein Embp facilitates colonization of surfaces through Velcro-like attachment to fibrillated fibronectin

  1. Nasar Khan
  2. Hüsnü Aslan
  3. Henning Büttner
  4. Holger Rohde
  5. Thaddeus Wayne Golbek
  6. Steven Joop Roeters
  7. Sander Woutersen
  8. Tobias Weidner
  9. Rikke Louise Meyer

  • Curated by eLife

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    Evaluation Summary:

    Staphylococcus epidermidis is a commensal that colonizes corneocytes of humans and other mammals. Colonization is crucial for many aspects of health including the development of our immune system and protection against invading pathogens. Embp is a long protein encoded by S. epidermidis on the cell surface that coats implanted foreign devices and host fibronectin, but this study shows that Embp does not bind to fibronectin in its soluble form, instead requiring surface binding to expose other epitopes to bind host fibronectin. This study uses atomic force microscopy to demonstrate these specific molecular interactions and their likely relevance to host-microbe interactions with prospects for pharmaceutical interventions.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)

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Abstract

Staphylococcus epidermidis causes some of the most hard-to-treat clinical infections by forming biofilms: Multicellular communities of bacteria encased in a protective matrix, supporting immune evasion and tolerance against antibiotics. Biofilms occur most commonly on medical implants, and a key event in implant colonization is the robust adherence to the surface, facilitated by interactions between bacterial surface proteins and host matrix components. S. epidermidis is equipped with a giant adhesive protein, extracellular matrix-binding protein (Embp), which facilitates bacterial interactions with surface-deposited, but not soluble fibronectin. The structural basis behind this selective binding process has remained obscure. Using a suite of single-cell and single-molecule analysis techniques, we show that S. epidermidis is capable of such distinction because Embp binds specifically to fibrillated fibronectin on surfaces, while ignoring globular fibronectin in solution. S. epidermidis adherence is critically dependent on multivalent interactions involving 50 fibronectin-binding repeats of Embp. This unusual, Velcro-like interaction proved critical for colonization of surfaces under high flow, making this newly identified attachment mechanism particularly relevant for colonization of intravascular devices, such as prosthetic heart valves or vascular grafts. Other biofilm-forming pathogens, such as Staphylococcus aureus, express homologs of Embp and likely deploy the same mechanism for surface colonization. Our results may open for a novel direction in efforts to combat devastating, biofilm-associated infections, as the development of implant materials that steer the conformation of adsorbed proteins is a much more manageable task than avoiding protein adsorption altogether.

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  1. Version published to 10.7554/elife.76164 on eLife
  2. eLife

    Evaluation Summary:

    Staphylococcus epidermidis is a commensal that colonizes corneocytes of humans and other mammals. Colonization is crucial for many aspects of health including the development of our immune system and protection against invading pathogens. Embp is a long protein encoded by S. epidermidis on the cell surface that coats implanted foreign devices and host fibronectin, but this study shows that Embp does not bind to fibronectin in its soluble form, instead requiring surface binding to expose other epitopes to bind host fibronectin. This study uses atomic force microscopy to demonstrate these specific molecular interactions and their likely relevance to host-microbe interactions with prospects for pharmaceutical interventions.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)

  3. eLife

    Reviewer #1 (Public Review):

    Staphylococcus epidermidis is a commensal that colonizes corneocytes of humans and other mammals. Colonization is crucial for many aspects of health including the development of our immune system and protection against invading pathogens causing skin and soft tissue infections including Staphylococcus aureus and Streptococcus pyogenes. However, certain strains of S. epidermidis, especially those that colonize hospitalized patients, have the ability to bind implantable foreign bodies (e.g. hip/knee/shoulder implants, catheters) and subsequently form biofilms. Biofilms are difficult to treat with antimicrobial agents, and, in many cases, the only method of treatment is to remove the device causing significant morbidity.

    Embp is a very long protein encoded by most strains of S. epidermidis that extends from the cell surface. It has been previously documented that Embp binds to the serum binding protein fibronectin, a glycoprotein that coats implanted foreign devices. Data from this manuscript documents that Embp does not bind to Fibronectin in its soluble form. However, once Fibronectin binds to a surface such as foreign bodies ( called the fibrillated form) other epitopes are exposed to which Embp binds. There are three major strengths of this manuscript. First, using elegant genetic techniques as well as advanced atomic force spectroscopy experiments, the investigators found that Embp binds only to fibrillated Fibronectin and not soluble Fibronectin. Second, they found that the large number of repeats associated with Embp function is similar to velcro where the strength of each interaction (between F- or FG- repeat and Fibronectin) is small but the additive nature of the interaction is significant. Lastly, they found using flow experiments that Embp functions in an environment with high shear stress similar to blood. This is a very exciting result and has a significant impact on the field, especially since other adhesins presumably facilitated adherence to Fibronectin in a low shear environment.

    In summation, the results suggest that new biomaterials can be designed to inhibit the formation of fibrillated fibrinogen. Indeed, studies in this manuscript document that Fibronectin primarily is found as a globular form when attached to compounds such as poly ethyl acrylate.

  4. eLife

    Reviewer #2 (Public Review):

    The objective of this study was to use atomic force microscopy to analyse the strength of the binding interaction between the fibronectin binding repeats of Embp and the fibrillary form of the host protein and to study bacterial adhesions under shear-stress.

    Strengths:

    -Using a surrogate host to study interactions between truncates of Embp with variable numbers of Fn binding repeats
    -Being able to attach globular Fn to a surface so that it either remains in the globular state or unfolds to reveal the buried binding domain
    -Studying bacterial attachment to immobilized Fn under flow conditions mimicking the blood stream
    -Applying atomic force microscopy to study the strength of binding between a single cell expressing Embp or a single molecule of Embp attached to the cantilever and surface-immobilized Fn allowing measurement of the strength of binding of individual and multiple Fn binding repeats

    Weaknesses:

    -The application of AFM is not discussed in the context of the extensive studies by the Dufrene group who have published many papers on the binding of staphylococcal adhesins to immobilized ligands.
    -The AFM analysis lacks detail, please add more to enable replication
    -The Dufrene group showed that some interactions are promoted by shear stress under flow conditions (for example SdrG and ClfA binding to fibrinogen). In these cases the force need to separate molecules is very strong and equivalent to that needed to break a covalent bond. It is not clear how the Embp-Fn bond responds to shear stress
    -There is no discussion or comparison of Fn binding by the well characterized FnBPs of S.aureus (used as a control in Figure 1) which bind to the type I repeats in the Fn N-terminus
    The authors have achieved their objectives and have advanced the state of knowledge about the interaction in an incremental fashion. It does not provide any major new insights. They have confirmed that Embp only binds the fibrillar form of Fn and they have shown that the strength of the interaction is proportional to the number of binding repeats in Embp truncates. This study confirms the value of force microscopy to studying bacterial adhesin-ligand interactions

  5. Version published to 10.1101/2021.05.31.446437v2 on bioRxiv
  6. Version published to 10.1101/2021.05.31.446437v1 on bioRxiv