Epitope Mapping Anti-ADAMTS13 Antibodies
Thomson, Jennifer A.
MetadataShow full item record
Thrombotic thrombocytopenic purpura (TTP) is a severe blood clotting disorder characterized by a pentad of clinical signs: hemolytic anemia, thrombocytopenia, fever, neurological dysfunction, and renal failure. The disorder is the result of a deficiency in the normal processing of the plasma glycoprotein von Willebrand Factor by the metalloprotease ADAMTS13. TTP can arise in one of two ways. Familial TTP is the result of a mutation in the ADAMTS13 gene, while acute TTP results from the presence of immunoglobulin-G autoantibodies directed against the protease. To further explore the role of antibodies in ADAMTS13 inhibition, this study examined the minimal peptide sequence recognized by a monoclonal antibody. Monoclonal antibodies raised in mice were used to immunoscreen random ADAMTS13 peptide fragments expressed by Lambda gt 11 bacteriophage. The ADAMTS13 fragments to which each monoclonal antibody bound were identified by sequencing the corresponding Lambda gt 11 cDNA, and the minimal linear peptide sequence necessary for antibody recognition and binding was determined. A minimal epitope of eight amino acids was identified in the metalloprotease domain and an epitope of 22 amino acids was identified in the disintegrin domain. This study showed that regions in both of these domains produced an immune response in mice. Future studies should first focus on further narrowing the epitopes for the monoclonal antibodies examined here. The project was also beneficial in testing a method that could be used to immunoscreen antibodies from human TTP patient plasma. Determining the regions of ADAMTS13 that produce an immune response in humans could be an essential first step in identifying a therapeutic target in humans with acute TTP.