This proposal will address the hypothesis that Porphyromonas gingivalis produces a newly identified angiosuppressive protein (ASP) that acts as a virulence factor by inhibiting angiogenesis and promoting chronic periodontal disease. Preliminary evidence from my laboratory indicates that the ASP is a low molecular weight, heat-labile protein which inhibits capillary tube formation and disrupts established capillary networks formed by human vascular endothelial cells cultured on Matrigel. Release of this virulence factor following bacterial lysis by the host's cellular defenses wold promote vascular destruction and create an anaerobic, hemin-rich environment conducive to bacterial growth.
The specific aims of this proposal are to: (1) isolate and purify the ASP from soluble sonic extracts of P. gingivalis by standard protein isolation methods, including molecular sieve and ion exchange chromatography, and preparative electrophoresis, (2) clone the ASP gene from a P. gingivalis genomic library by standard methods (PCR or expression cloning), (3) prepare ASP antisera for screening recombinant plasmid-transformed clones, and (4) clearly define the morphometric parameters for measuring the effects of the ASP on in vitro capillary tube formation.
