Identification and biochemical characterization of two novel collagen binding MSCRAMMs of Bacillus anthracis

Cell wall-anchored proteins play critical roles in the pathogenesis of infections caused by Gram-positive bacteria. Through the analysis of the genome of Bacillus anthracis Ames strain, we identified two novel putative cell wall-anchored proteins, BA0871 and BA5258, which have sequence homology to CNA, a cell wall-anchored collagen adhesin of Staphylococcus aureus. The two proteins have similar domain organization to that of CNA, with typical signal peptide sequences, a non-repetitive A region followed by repeats, and a characteristic cell wall-anchoring region. They are expressed on the surface of B. anthracis. The A regions of the two proteins were predicted to adopt similar structural folds as CNA. Circular dichroisnt analysis of the recombinant A regions of the two proteins (rBA0871A and rBA5258A) indicate that their secondary structure compositions are similar to those of the A regions of CNA and other cell wall-anchored adhesins. We demonstrate through solid phase binding assays and surface plasmon resonance analyses that rBA0871A and rBA5258A specifically bound type I collagen in a dose-dependent and saturable manner. Their dissociation constants (K_D) for collagen are 1.6-3.2 μM for rBA0871A and 0.6-0.9 μM for rBA5258A, respectively. We further demonstrate that BA0871 and BA5258 can mediate cell attachment to collagen when expressed on the surface of a heterologous host bacterium. To our knowledge these are the first two adhesins of B. anthracis described, which may have important implications for our understanding of the pathogenic mechanisms explored by this organism.