Exclusion of two major areas on thyroid peroxidase from the immunodominant region containing the conformational epitopes recognized by human autoantibodies.
We have used a chimeric molecule between thyroid peroxidase (TPO) and myeloperoxidase (MPO) as well as new information on the three-dimensional structure of MPO to refine further our understanding of the location of the TPO-immunodominant region recognized by TPO autoantibodies in patients' sera. In TPO-MPO chimera A, the amino-terminal 146 amino acids of MPO substitute for the amino-terminal 121 amino acids of TPO. We performed fluorescence-activated cell sorter analysis of Chinese hamster ovary cells expressing TPO-MPO-A on their surface using four monoclonal human autoantibody F(ab) (WR1.7, TR1.8, TR1.9, and SP1.4) that define the immunodominant region. All four F(ab) recognized the TPO-MPO-A chimeric molecule to the same extent. In a second approach to refine the location on the TPO-immunodominant region, we compared the ability of the TPO autoantibody F(ab) to inhibit the binding of serum autoantibodies to the monomeric and dimeric forms of human TPO. The F(ab) inhibited equally (approximately 80%) the binding to the TPO monomer and dimer by autoantibodies in the sera of six individual patients. The present observations exclude two major regions of TPO from the autoantibody-immunodominant region, namely the amino-terminal 121 amino acids of the TPO extracellular domain and the contact region between the two TPO monomers. These findings together with previous data on the Mab47/C21 region of TPO and the recently elucidated 3-dimensional structure of highly homologous MPO, narrow, by a process of exclusion, the site on TPO comprising the immunodominant region. The data provide further support for the thesis, still controversial, that the majority of TPO autoantibodies recognize the native molecule.
Nishikawa
T
T
Thyroid Molecular Biology Unit, Veterans Administration Medical Center, San Francisco, California.
Rapoport
B
B
McLachlan
S M
SM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology; D001323:Autoantibodies / Q000276:immunology*; D016466:CHO Cells; D006224:Cricetinae; D000939:Epitopes / Q000737:chemistry* / Q000276:immunology; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D007140:Immunoglobulin Fab Fragments / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D007453:Iodide Peroxidase / Q000737:chemistry* / Q000276:immunology*; D046911:Macromolecular Substances; D008958:Models, Molecular; D015394:Molecular Structure; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments; D046911:Macromolecular Substances; D011993:Recombinant Fusion Proteins; D007453:Iodide Peroxidase