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 targets of humoral autoimmunity in patients with insulin-dependant diabetes mellitus (IDDM). In an effort to define the epitopes recognized by autoantibodies against IA-2, we generated five human mAbs (hAbs) from peripheral B lymphocytes isolated from patients most of whom had been recently diagnosed for IDDM. Determination and fine mapping of the critical regions for autoantibody binding was performed by RIA using mutant and chimeric constructs of IA-2- and IA-2beta-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 cross-reacted with the related IA-2beta PTP-like domain (IA-2beta aa 741-1033). A further PTP-like domain specific hAb required the entire PTP-like domain (aa 687-979) for binding, but critical amino acids clustered in the N-terminal 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 associated with IDDM. The presence of somatic mutations in both heavy and light chain genes and the high affinity or this Ab suggest that the immune response to IA-2 is Ag driven.
Kolm-Litty
V
V
Institut für Diabetesforschung, Munich, Germany.
Berlo
S
S
Bonifacio
E
E
Bearzatto
M
M
Engel
A M
AM
Christie
M
M
Ziegler
A G
AG
Wild
T
T
Endl
J
J
D000328:Adult; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000145:classification / Q000302:isolation & purification*; D001323:Autoantibodies / Q000096:biosynthesis; D001324:Autoantigens / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D001666:Binding Sites, Antibody / Q000235:genetics; D001667:Binding, Competitive / Q000235:genetics / Q000276:immunology; D002461:Cell Line, Transformed; D002648:Child; D003922:Diabetes Mellitus, Type 1 / Q000201:enzymology / Q000235:genetics / Q000276:immunology*; D018604:Epitope Mapping; D005822:Genetic Vectors / Q000138:chemical synthesis / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000096:biosynthesis; D007135:Immunoglobulin Variable Region / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008875:Middle Aged; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000276:immunology; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000276:immunology*; D054635:Receptor-Like Protein Tyrosine Phosphatases, Class 8; D017421:Sequence Analysis; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D001324:Autoantigens; D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region; D008565:Membrane Proteins; C516862:PTPRN protein, human; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; C516863:Ptprn protein, mouse; D054635:Receptor-Like Protein Tyrosine Phosphatases, Class 8