Structure and function of staphylococcal surface proteins involved in biofilm growth and virulence

A major focus of our laboratory is to broadly understand the molecular mechanisms by which cell wall- anchored (CWA) proteins on the surface of Staphylococcus epidermidis and S. aureus promote biofilm formation and virulence. Staphylococcal biofilms are highly adhesive and cohesive communities of surface-

adherent bacteria that are highly resistant to antibiotic action and host immune responses, often resulting in recalcitrant infections. Specifically, the research will focus on the large, multi-domain CWA protein Aap from S. epidermidis, its ortholog SasG from S. aureus, and another large S. aureus CWA protein called SasC; each of

these is known to mediate homophilic interactions that promote intercellular adhesion. Mechanisms of both reversible self-assembly and nucleation of functional amyloid fibrils will be studied in order to identify avenues for therapeutic intervention to prevent biofilm formation. In addition, the small, secreted S. epidermidis protein

SBP and its ortholog from S. aureus will be investigated in terms of their ability to interact with Aap (and SasG) to facilitate assembly and potentially trigger liquid-liquid phase separation of macromolecular components in the biofilm matrix. Finally, the S. aureus protein SasX, implicated in the spread of a recent epidemic of

methicillin-resistant S. aureus, is also cell wall-anchored but unlike the other CWA proteins is a small, intrinsically disordered protein. SasX will be assessed for its ability to interact with other staphylococcal surface proteins as well as keratinocyte ligands, given its demonstrated role in promoting biofilm formation as well as

facilitating adhesion to epithelial cells and nasal colonization. In addition to structural and biophysical studies of these proteins, quantitative analysis of biofilm morphology and mechanical properties using confocal microscopy, rheometry, and force spectroscopy will provide complementary insights in a biological context.

Over the next five years, our goal is to provide a structural and functional view of how the most important surface proteins in S. aureus and S. epidermidis physically hold the cells together in biofilms and to understand how those protein-protein interactions mechanically strengthen the biofilm. By understanding these interactions

at the molecular level, our goal is to identify sites of vulnerability that will allow the design of therapeutic approaches to target the ability of these bacteria to form biofilms, rendering them susceptible to a broader range of antimicrobial agents.