High Affinity Binding of Escherichia coli Cytotoxic Necrotizing Factor 1 (CNF1) to Lu/BCAM Adhesion Glycoprotein
نویسندگان
چکیده
The protein toxin Cytotoxic Necrotizing Factor 1 (CNF1) is a major virulence factor of pathogenic Escherichia coli strains. It belongs to a family of single chain AB-toxins, which enter mammalian cells by receptor-mediated endocytosis. Recently, we identified the Lutheran (Lu) adhesion glycoprotein/basal cell adhesion molecule (BCAM) as a cellular receptor for CNF1. Here, we identified the Ig-like domain 2 of Lu/BCAM as main interaction site of the toxin by direct protein-protein interaction and competition studies. Using surface plasmon resonance, we showed a high affinity CNF-Lu/BCAM interaction with a KD of 2.8 nM. Furthermore, we performed small-angle X-ray scattering to define the molecular envelope of the Lu/BCAM-CNF1 complex, suggesting a 6:1 ratio of Lu/BCAM to CNF1 in the receptor-toxin complex. This study leads to a deeper understanding of the interaction between CNF1 and Lu/BCAM, and presents novel opportunities for the development of future anti-toxin strategies.
منابع مشابه
Lu/BCAM Adhesion Glycoprotein Is a Receptor for Escherichia coli Cytotoxic Necrotizing Factor 1 (CNF1)
The Cytotoxic Necrotizing Factor 1 (CNF1) is a protein toxin which is a major virulence factor of pathogenic Escherichia coli strains. Here, we identified the Lutheran (Lu) adhesion glycoprotein/basal cell adhesion molecule (BCAM) as cellular receptor for CNF1 by co-precipitation of cell surface molecules with tagged toxin. The CNF1-Lu/BCAM interaction was verified by direct protein-protein int...
متن کاملAnalysing the Structural Effect of Point Mutations of Cytotoxic Necrotizing Factor 1 (CNF1) on Lu/BCAM Adhesion Glycoprotein Association
Cytotoxic Necrotizing Factor 1 (CNF1) was identified in 1983 as a protein toxin produced by certain pathogenic strains of Escherichia coli. Since then, numerous studies have investigated its particularities. For instance, it is associated with the single chain AB-toxin family, and can be divided into different functional and structural domains, e.g., catalytic and transmembrane domain and inter...
متن کاملIncreased Cell-Matrix Adhesion upon Constitutive Activation of Rho Proteins by Cytotoxic Necrotizing Factors from E. Coli and Y. Pseudotuberculosis
Cytotoxic necrotizing factors (CNFs) encompass a class of autotransporter toxins produced by uropathogenic E. coli (CNF1) or Y. pseudotuberculosis (CNFy). CNF toxins deamidate and thereby constitutively activate RhoA, Rac1, and Cdc42. In this study, the effects of CNF1 on cell-matrix adhesion are analysed using functional cell-adhesion assays. CNF1 strongly increased cell-matrix binding of susp...
متن کاملEscherichia coli cytotoxic necrotizing factor 1 inhibits intestinal epithelial wound healing in vitro after mechanical injury.
Cytotoxic necrotizing factor type 1 (CNF1) from Escherichia coli activates the small GTP-binding proteins of the Rho family (Rho, Rac, and Cdc42) by catalyzing their deamidation at a specific glutamine residue. Since RhoA, Rac, and Cdc42 play a pivotal role in cell migration during the early phase of wound repair, we investigated whether CNF1 was able to interfere with wound healing in intestin...
متن کاملThe p21 Rho-activating toxin cytotoxic necrotizing factor 1 is endocytosed by a clathrin-independent mechanism and enters the cytosol by an acidic-dependent membrane translocation step.
Cytotoxic necrotizing factor 1 (CNF1), a protein produced by pathogenic strains of Escherichia coli, activates the p21 Rho-GTP-binding protein, inducing a profound reorganization of the actin cytoskeleton. CNF1 binds to its cell surface receptor on HEp-2 cells with high affinity (K(d) = 20 pM). In HEp-2 cells the action of CNF1 is not blocked in the presence of filipin, a drug described to redu...
متن کامل