Single Molecular Force Sensing Reveals Fibronectin-Specific Binding Epitopes of Bacterial Curli Fimbriae

Abstract

Curli are functional microbial amyloids that are major components of the complex extracellular matrix produced by Enterobacteriaceae. They play important functions in biofilm formation, bacterial invasion into host cells, and pathological processes which lead to numerous infections and autoimmune diseases. Curli fibers interact with host cell multi-domain adhesive glycoproteins, such as laminin and fibronectin. Several groups have investigated the structure of curli fimbriae, as well as their mechanical properties and adhesion mechanisms. However, the specificity and exact mechanism of curli-mediated bacterial binding to cells remains obscure. Here, single-molecule force spectroscopy reveals dynamic and statistical information about the formation of molecular bonds involved in the adhesion of curliated bacteria to fibronectin. We show that a specific interaction between the bacterial curli protein CsgA and the RGD domain of fibronectin leads to tight binding. Via single-cell force spectroscopy we measured distinct interaction forces on the surface between the isolated CsgA protein or the bacteria-producing CsgA curli and full length fibronectin, fibronectin domain III, and RGD peptide, respectively. These experiments demonstrate that the forces required to break adhesion are quantized and reflect formation of a dense collective network. These networks comprise at least 20 specific molecular bonds that lead to tight bacterial binding. Our results underline the significance of amyloid formation on bacterial surfaces as multi-bond structural components in curli-mediated bacterial adhesion to fibronectin.