Functional, structural and evolutionary studies on a family of bacterial surface proteins

Sammanfattning: This thesis is concerned with a family of bacterial surface proteins expressed by different strains of the anaerobic Gram-positive, commensal and human pathogen Peptostreptococcus magnus. These proteins have been studied with respect to their role in virulence, function, structure, and evolution. Strains isolated from patients with vaginosis have previously been shown to be associated with expression of the immunoglobulin (Ig) light chain-binding protein L. Here it was shown that binding of human serum albumin (HSA) is associated with localised suppurative infections; such as wound infections and abscesses. However, commensal strains bound neither IgG nor HSA. Studies of the binding of protein L to Ig of various mammalian species other than human, showed activity in 12 out of 23 Ig screened, mainly from primates and rodents, stressing the versatility of protein L as a immunochemical tool. Protein PAB, a novel HSA-binding protein was isolated from P. magnus and characterised. The gene sequence revealed homology of the HSA-binding domain to the HSA-binding regions of the group G streptococcal protein G. This was taken to be the first contemporary example of module shuffling. The HSA-binding motif (the GA module) of 45 amino acid residues was cloned and expressed in E. coli and the secondary structure and global fold were determined by NMR and shown to be an antiparallel three-helix-bundle. A second HSA-binding protein, protein urPAB, was isolated from a strain showing less binding activity. This protein was the presumptive predecessor of protein PAB as it lacked the shuffled GA module. Proteins PAB, urPAB, L and another previously characterised protein L-like protein, shared common structural motifs, but differed in types and numbers of functional modules. They were taken to be homologues, making up a bacterial surface protein family. A detailed analysis of the corresponding genes revealed that a consensus sequence (recer) of 15 nucleotides flanked most of the various modules. The recer sequence was thought to play a role as a site for recombination in the evolution of this protein family via module shuffling. It was also shown that bacterial strains of different species, P. magnus and group G streptococci, that have incorporated the same HSA-binding module into their surface proteins exhibit substantially higher growth rates when the growth medium is supplemented with HSA. This finding also has implications for the function of other bacterial albumin-binding proteins, and suggests that the bacteria expressing this property experience a selective advantage.