2 DISTINCT GLUTAMIC-ACID DECARBOXYLASE AUTOANTIBODY SPECIFICITIES IN IDDM TARGET DIFFERENT EPITOPES

Although most individuals with insulin-dependent diabetes mellitus (ID DM) have autoantibodies to glutamic acid decarboxylase (GAD), antibodi es to GAD are also present in some individuals with a low risk of deve loping diabetes. The GAD autoantibodies of IDDM are specific for the G AD(65) isoform, do not bind denatured GAD protein, and target epitope( s) dependent on conformation of the protein. However, the IDDM epitope s have been difficult to further define because the antibodies do not bind GAD protein fragments or synthetic peptides. Since the GAD(67) is oform is highly homologous to GAD(65) but is usually not a target of t he GAD autoantibodies in IDDM sera, we created six GAH(65)/GAD(67) chi meric proteins to maintain the overall GAD protein conformation and us ed these chimeric proteins to map conformation-dependent epitopes of G AD(65) targeted by IDDM sera. We find that the GAD binding present in most IDDM sera (n = 11 of 12) is composed of two distinct GAD antibody specificities that target different conformation-dependent regions of the GAD(65) protein, one that is located between amino acids 240 and 435 (termed IDDM-E1) and one that is located between amino acids 451 a nd 570 (termed IDDM-E2). One IDDM serum (n = 1 of 12) bound only the I DDM-E1 region. Identification of epitopes targeted by IDDM sera may al low one to distinguish between GAD antibody-positive individuals at hi gh and low risk of developing IDDM and to determine if differences in the autoimmune repertoire directed at GAD are present. The chimeric GA D(65)/GAD(67) proteins may also be useful in designing GAD assays spec ific for IDDM.