Extreme Sequence Divergence but Conserved Ligand-Binding Specificity in Streptococcus pyogenes M Protein

Abstract

Many pathogenic microorganisms evade host immunity through extensive sequence variability in a protein region targeted by protective antibodies. In spite of the sequence variability, a variable region commonly retains an important ligand-binding function, reflected in the presence of a highly conserved sequence motif. Here, we analyze the limits of sequence divergence in a ligand-binding region by characterizing the hypervariable region (HVR) of Streptococcus pyogenes M protein. Our studies were focused on HVRs that bind the human complement regulator C4b-binding protein (C4BP), a ligand that confers phagocytosis resistance. A previous comparison of C4BP-binding HVRs identified residue identities that could be part of a binding motif, but the extended analysis reported here shows that no residue identities remain when additional C4BP-binding HVRs are included. Characterization of the HVR in the M22 protein indicated that two relatively conserved Leu residues are essential for C4BP binding, but these residues are probably core residues in a coiled-coil, implying that they do not directly contribute to binding. In contrast, substitution of either of two relatively conserved Glu residues, predicted to be solvent-exposed, had no effect on C4BP binding, although each of these changes had a major effect on the antigenic properties of the HVR. Together, these findings show that HVRs of M proteins have an extraordinary capacity for sequence divergence and antigenic variability while retaining a specific ligand-binding function. Many pathogens have evolved mechanisms to evade host immunity. In one such mechanism, the sequence of a surface protein varies among different strains of a pathogen. This sequence variability represents an apparent paradox, because the variable protein must retain an important function. The authors studied this problem in Streptococcus pyogenes, a major human pathogen. The surface-localized M protein of this bacterium varies extensively in sequence between bacterial strains, allowing immune escape. Nevertheless, the most variable part of the M protein commonly binds a human plasma protein. By hijacking this human protein the bacteria evade attack by complement an important part of the innate immune system. Comparison of the ligand-binding region in different M proteins showed that these regions lack a shared amino acid sequence motif. Thus, a variable protein can retain a ligand-binding function in the absence of a conserved binding motif. Evidence is also presented that a single amino acid change in the variable region may cause a major antigenic change, providing a selective advantage for the bacteria. Together, these data bear witness to the extraordinary ability of pathogens to escape host immunity, without losing ability to cause disease.