Specific interactions between F1 adhesin of Streptococcus pyogenes and N-terminal modules of fibronectin

Protein F1 is a surface protein of Streptococcus pyogenes that mediates hig h affinity binding to fibronectin (Fn) and facilitates S. pyogenes adherenc e and penetration into cells. The smallest portion of F1 known to retain th e full binding potential of the intact protein is a stretch of 49 amino aci ds known as the functional upstream domain (FUD). Synthetic and recombinant versions of FUD were labeled with fluorescein isothiocyanate and used in f luorescence anisotropy experiments. These probes bound to Fn or the 70-kDa fragment of Fn with dissociation constants of 8-30 nM. Removal of the N-ter minal seven residues of FUD did not cause a change in binding affinity. Fur ther N- or C-terminal truncations resulted in complete loss of binding acti vity. Analysis of recombinant versions of the 70-kDa fragment that lacked o ne or several type I modules indicates that residues 1-7 of the 49-mer bind to type I modules I1 and I2 of the 27-kDa subfragment and the C-terminal r esidues bind to modules I4 and I5. Fluorescein isothiocyanate-labeled 49-me r also bound with lower affinity to large Fn fragments that lack the five t ype I modules of the 27-kDa fragment but contain the other seven type I mod ules of Fn. These results indicate that, although FUD has a general affinit y for type I modules, high affinity binding of FUD to Fn is mediated by spe cific interactions with N-terminal type I modules.