Autoantibodies to IA-2 and IA-2ß in Insulin-Dependent Diabetes Mellitus Recognize Conformational Epitopes
Hong Xie; Baowei Zhang; Yasunobu Matsumoto; Qing Li; Abner 1. Notkins; Michael S. Lan
IA-2 and IA-2ß are major autoantigens in insulin-dependent diabetes mellitus (IDDM) and the precursors, respectively, of a 40 and 37-kDa tryptic fragment that reacts with IDDM sera. In the present study, by amino acid sequencing of recombinant IA-2 and IA-2ß, we determined the tryptic cleavage sites involved in the generation of these fragments. Both cleavage sites are immediately after an arginine residue at position 653 for IA-2 and position 679 for IA-2ß. The resulting tryptic fragments are 326 and 307 amino acids in length and retain their ability to react with IDDM sera. In contrast to IA-2 and IA-2ß, other members of the protein tyrosine phosphatase (PTP) family (i.e., RPTPK, RPTPp, NU-3, SHP, and 3CH134) are completely susceptible to
digestion by trypsin. Sequence analysis revealed five conserved cysteine residues in IA-2 and IA-2ß that are not present in other PTPs. Reduction and alkylation of IA-2 and IA-2ß recombinant proteins resulted in loss of both resistance to digestion by trypsin and reactivity with autoantibodies in IDDM sera. It is concluded that disulfide bond formation plays a critical role in the
maintenance of antigenic structure and that the autoantibodies to IA-2 / IA-2ß in IDDM sera recognize conformational epitopes. The journal of Immunology, 1997, 159: 3662-3667.