The multiple and often serotype-specific functions of M proteins are generally poorly understood in molecular terms, arguably due to a lack of structural information. All available evidence supports the idea of M proteins adopting an elongated dimeric, parallel coiled-coil conformation. This makes M proteins difficult targets for conventional structural biology. We combined EPR and NMR spectroscopy to map the hypervariable region (HVR) in full length M3 protein in solution and derive a structural model. M3-HVR was found to form a well-defined folded structure that deviates from coiled coil topology. This fold presents collagen-binding motifs associated with rheumatic fever in a structural context that depends on the dimeric state of the protein and is required for binding activity. Using triple-helical peptide libraries we identified M3-binding regions within the triple-helical domain of collagen II. The M protein:collagen interaction was found to mediate biofilm formation of M3 clinical isolates.
Our study reveals a surprising structural complexity of the M3 protein with implications for its function in rheumatic fever and biofilms. It also suggests that M proteins may be structurally more diverse than assumed.