Genetic dissection of the Streptococcus pyogenes M1 protein: regions involved in fibronectin binding and intracellular invasion

Entry of serotype M1 Streptococcus pyogenes into host cells depends on bind ing of the host glycoprotein fibronectin (Fn) by the bacterial M1 protein. The present study was undertaken to localize the Fn binding region in M1 an d assess other potential functions of M1. A set of recombinant M1 protein f ragments were assayed for their capacities to bind Fn and inhibit ingestion of streptococci by epithelial cells. M1 protein, M6 protein and internally -deleted derivatives of M1 were expressed on the surface of Lactococcus lac tis. Lactococci that expressed M1 or M6 protein bound Fn and were efficient ly taken up by epithelial cells. Deletion of both the N-terminal A and B re peats regions of M1 abrogated Fn binding and intracellular invasion. Deleti on of either the A domain (M1 DeltaA) or B repeats (M1 DeltaB) significantl y reduced, but did not completely eliminate, Fn binding indicating that M1 protein may possess two independent Fn binding sites. Fn binding by the M1 DeltaA or M1 DeltaB proteins was insufficient for efficient invasion, howev er, suggesting that M protein binding alters the structure of Fn that, in t urn, affects the interaction between Fn and epithelial cells. Although expr ession of M1, M6 or M1 AB proteins led to aggregation of lactococcal cells, aggregation did not significantly contribute to invasion efficiency. (C) 2 001 Academic Press.