Molecular interactions: oral bacteria & matrix proteins

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Gram-negative bacteria are associated with periodontal diseases, which are a group of chronic inflammatory diseases of the gingiva and the supporting structures of the periodontium. Actinobacillus actinomycetemcomitans (Aa) is a Gram-negative, facultative coccobacillus that colonizes the human oral cavity and upper respiratory tract. This bacterium is closely associated with periodontitis in young individuals and with cases of adult periodontitis. This pathogen has been associated with other serious human infections such as endocarditis, soft tissue abscesses, and more recently cardiovascular disease. Although the periodontium is believed to be the source of these extraoral infections, little is known about the tropisms used by Aa to maintain itself within the oral cavity and to infiltrate and disseminate in tissues. Pathogens have evolved diverse strategies to be successful in colonization of the host tissue. A common theme amongst these pathogens is the ability to initiate infection by adhesion to specific host macromolecules under stringent or hostile conditions. These molecules include proteins that are secreted by host cells that form the extracellular matrix. This matrix is usually composed of collagen and specific noncollagenous proteins, e.g. fibronectin. The major protein found in the periodontium is collagen and we have demonstrated that Aa binds to both collagen and fibronectin. In this proposal, we plan to identify the genes coding for matrix binding proteins and determine the amino acid sequences of these proteins required for binding. It is our hypothesis that the synthesis of matrix binding proteins is involved in Aa colonization of the periodontal pocket and underlying tissues. In order to realize these goals, we propose to 1) isolate the genes coding for collagen and fibronectin binding proteins by constructing a transposon mutagenesis and a phage display library; 2) determine the gene sequences and generate isogenic mutants; 3) determine the regions(s) of the protein essential for binding activity; 4) determine the immunoreactivity of LJP patient sera to these proteins. This information can be used for the development of therapeutic agents of vaccines for periodontal disease.