Structure-Function Studies of Streptococcus Fibrillar Adhesin CshA

Many species of bacteria express cell-surface adhesins that promote colonisation and infection through host receptor recognition. Streptococcus gordonii is a commensal primary coloniser of the oral cavity, but is also implicated as a leading cause of infective endocarditis. S. gordonii expresses several cell-surface proteins that enable the bacteria to colonise a variety of sites throughout the human body. One such protein is CshA (259 kDa). CshA forms fibrils that extend away from the bacterial cell surface (~60 nm long), interact with cellular fibronectin, and mediate co-aggregation with other oral microorganisms in the formation of biofilm communities. However, the molecular mechanisms that underpin these interactions are unknown. Objectives: The aims of this study were to obtain detailed structural information about CshA, and to define its functional regions in S. gordonii colonisation and pathogenesis. Methods: Based upon structural predictions, CshA was divided into distinct regions. These regions and combinations thereof were then expressed and purified as recombinant proteins in Escherichia coli, and subjected to structural analyses, including X-ray crystallography, SAXS and NMR. Results: The N-terminal portion of CshA was shown to be disordered, whilst the rest is mainly beta-sheet. There was also evidence for aggregation of segments of the non-repetitive region, which may assist in fibril formation. Conclusions: These initial data imply that CshA has a novel structure not previously reported for bacterial adhesins. CshA-like polypeptides are expressed by a wide range of streptococci. Thus by determining the structural and functional properties of CshA, these studies may provide insights into novel mechanisms of streptococcal colonisation and pathogenesis.