Design of aStreptococcus pyogenesM protein immunogen to elicit M type cross-reactivity

Coiled coil-forming M proteins of the widespread and potentially deadly bacterial pathogen Streptococcus pyogenes (Strep A) are immunodominant targets of opsonizing antibodies. However, antigenic sequence variability into >220 M types, as defined by the M protein hypervariable region (HVR), has been considered to limit its utility as a vaccine immunogen due to type-specificity in the antibody response. Surprisingly, a multi-HVR immunogen in clinical vaccine trials elicited M type cross-reactivity. The basis for this cross-reactivity is unknown but may be due in part to antibody recognition of a three-dimensional (3D) pattern conserved in many M protein HVRs that confers binding to human C4b-binding protein (C4BP). To test this hypothesis, we asked whether a single M protein immunogen carrying the 3D pattern would elicit cross-reactivity against other M types carrying the 3D pattern. We found that a 34-amino acid sequence of M2 protein bearing the 3D pattern retained full C4BP-binding capacity when fused to a coiled coil-stabilizing sequence from GCN4. This immunogen, called M2G, elicited cross-reactive antibodies against a number of M types that carry the 3D pattern but not against those that lack the 3D pattern. The M2G antiserum recognized M proteins as displayed natively on the Strep A surface, and promoted the opsonophagocytic killing of Strep A strains expressing these M proteins. As C4BP-binding is a conserved virulence trait of Strep A, targeting the 3D pattern may prove advantageous in vaccine design.