Human monoclonal antibodies isolated from type I diabetes patients define multiple epitopes in the protein tyrosine phosphatase-like IA-2 antigen

Protein tyrosine phosphatase-like IA-2 autoantigen is one of the major targ ets of humoral autoimmunity in patients with insulin-dependant diabetes mel litus (IDDM). In an effort to define the epitopes recognized by autoantibod ies against IA-2, we generated five human mAbs (hAbs) from peripheral B lym phocytes isolated from patients most of whom had been recently diagnosed fo r IDDM. Determination and fine mapping of the critical regions for autoanti body binding was performed by RIA using mutant and chimeric constructs of I A-2- and IA-2 beta-regions, Four of the five IgG autoantibodies recognized distinct epitopes within the protein tyrosine phosphatase (PTP)-like domain of IA-2, The minimal region required for binding by three of the PTP-like domain-specific hAbs could be located to aa 777-979, Two of these hAbs cros s-reacted with the related IA-2 beta PTP-like domain (1A-2 beta aa 741-1033 ), A further PTP-like domain specific hAb required the entire PTP-like doma in (aa 687-979) for binding, but critical amino acids clustered in the N-te rminal region 687-777, An additional epitope could be localized within the juxtamembrane domain (aa 603-779), In competition experiments, the epitope recognized by one of the hAbs was shown to be targeted by 10 of 14 anti-IA- 2-positive sera. Nucleotide sequence analysis of this hAb revealed that it used a V-H germline gene (DP-71) preferably expressed in autoantibodies ass ociated with IDDM. The presence of somatic mutations in both heavy and ligh t chain genes and the high affinity or this Ab suggest that the immune resp onse to IA-2 is Ag driven.