Functional Analysis of the Staphylococcus aureus Collagen Adhesin B Domain

Abstract

The Staphylococcus aureus collagen adhesin (CNA) occurs in at least four forms that differ in the number (one, two, three, or four) of B domains. The B domains contain 187 amino acids and are located between the domains that anchor CNA to the cell envelope and the ligand-binding A domain. To determine whether a B domain is required for functional expression of CNA, we cloned the 2Bcna gene from S. aureus strain Phillips and then eliminated both B domains by overlapping PCR. The absence of a B domain did not affect processing of the collagen adhesin to the cell surface or the ability to bind collagen. Based on our recent demonstration that the capsule can mask CNA on the surface of S. aureus cells (A. F. Gillaspy et al., Infect. Immun. 66:3170–3178, 1998), we also investigated the possibility that multiple B domains can extend the ligand-binding A domain outward from the cell surface and thereby overcome the inhibitory effect of the capsule. Specifically, we cloned the naturally occurring 4B CNA variant from S. aureus UAMS-639 and, by successive elimination of B domains, generated 1, 2, and 3B variants that are isogenic with respect to the 4B clone. After introducing each variant into microencapsulated and heavily encapsulated strains of S. aureus and growing cells under conditions known to affect capsule production (e.g., growth on Columbia agar), we correlated capsule production with exposure of CNA on the cell surface and the ability to bind collagen. Under no circumstance was the masking effect of the capsule reduced by the presence of multiple B domains. These results indicate that the B domains do not extend the ligand-binding A domain outward in a fashion that can overcome the inhibition of collagen binding associated with capsule production.

Collagen‐binding proteins of Streptococcus mutans and related streptococci

“…It has been suggested that the S. mutans Cnm and Cbm use a similar mechanism to bind collagen (Kang et al ., ), although the crystal structure for either protein has yet to be resolved. The B domain of Cna, composed of one to four repeating subunits of 23 kDa, does not alter the capacity of the A domain to interact with collagen (Rich et al ., ; Snodgrass et al ., ). Although the Cna B‐domain does not appear to be glycosylated, the B domain of Cnm undergoes O ‐glycosylation, a process that was shown to confer proteolytic stability and, possibly, function (Avilés‐Reyes et al ., ).…”

Isopeptide bond in collagen- and fibrinogen-binding MSCRAMMs

“…Concerning the repetitive B-region in collagen-binding MSCRAMMs, so far, the crystal structure consisting of two (PDB ID 1D2O) and four B-repeats (PDB ID 1D2P) (we considered one CnaB fold domain as one repeat) are available for collagen-binding Cna of S. aureus (Deivanayagam et al 2000). Despite the availability of these structures, the function of this Cna B-region is still unclear and is hypothesized to be functioning independently from the Cna A-region (Snodgrass et al 1999). Kang et al (2007) reported the presence of a putative isopeptide bond in the structure of Cna B-repeat.…”