Lyme disease, caused by the tick-borne spirochete Borrelia burgdorferi, is a chronic multisystemic illness. We have determined that B. burgdorferi bind to proteoglycans present on host cells and in extracellular matrix, a property likely to contribute to the spirochete's ability to colonize host tissues. A diverse collection of infectious Borrelia strains were shown to express this activity, as measured by the ability attach to dextran sulfate and heparin, and to agglutinate rabbit erythrocytes. In order to characterize the Borrelia heparin-binding hemagglutinin(s), the following goals will be pursued: (1) Purify the Borrelia heparin-binding hemagglutinin(s). The heparin- binding hemagglutinin will be purified from detergent extracts of B. burgdorferi, using carbohydrate affinity chromatography and other standard biochemical techniques. (2) Isolate monoclonal antibodies that block hemagglutination activity. Mice will be immunized with fractions enriched for the hemagglutinin(s). Hybridoma supernatants will be screened for inhibition of hemagglutination by sonicated bacterial extracts. (3) Isolate molecular clones that express the Borrelia hemagglutinin(s). Amino acid sequences derived from the purified hemagglutinin(s) will be used to generate oligonucleotide probes that will identify molecular clones carrying the hemagglutinin gene(s). In addition, clones that express the hemagglutinin(s) will by isolated from a B. burgdorferi expression library by reactivity with monoclonal antibodies. (4) Identify the heparin-binding domain of the Borrelia hemagglutinin(s). The epitopes recognized by the blocking monoclonal antibodies raised in Aim 2 will be mapped using fragments of the hemagglutinin(s). These fragments will also be tested for hemagglutination and proteoglycan binding activity. The bacterial determinants that mediate interactions of the spirochete with host molecules are likely to be important virulence determinants in Lyme disease. Characterization of these molecules will provide insights into the pathogenic mechanisms of this bacterium, and may provide novel strategies for therapeutic intervention.
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