Streptococcus mutans Collagen-Binding Protein Cnm Is a Multifunctional Adhesin: A Structural Investigation.

Abstract

The collagen-binding adhesin Cnm is a known virulence factor of Streptococcus mutans. It is present in specific serotypes (mostly e, f, and k strains) of S. mutans and belongs to the LPXTG family of cell wall-anchored surface adhesins. Here, we report the crystal structure of the collagen-binding N₂ domain of S. mutans Cnm. Using the Staphylococcus aureus collagen-binding protein Cna, which shares high sequence and structural homology with Cnm, we modeled collagen binding to S. mutans Cnm. The comparative analysis identified three conserved collagen-binding residues (Y176, F192, N194) and four equivalent residues that are different in their composition (D224, T226, S232, M276). This study also discovered the multifunctional attributes of this protein, where Cnm-FL, Cnm-N_12, and the individual domains of Cnm-N₁ and Cnm-N₂ adhere with high affinity to the scavenger receptor cysteine-rich (SRCR) domains of glycoprotein 340 (Gp340). Protein-protein docking of Cnm-N₂ and SRCR₁ showed the possibility of a shared binding site at the collagen-binding interface of Cnm-N₂. Furthermore, competition experiments using collagen and SRCR₁₂₃ with Cnm-N₂, Cnm-N_12, and Cnm-FL constructs confirmed that collagen and SRCR₁ share a binding site. Subsequent alanine substitution mutagenesis of the predicted collagen-binding residues validated our modeling results, confirming that Y176 and F192 are important residues for collagen and SRCR/Gp340 binding.