Analysis of a conformational B cell epitope of human thyroid peroxidase: identification of a tyrosine residue at a strategic location for immunodominance.
Thyroid peroxidase (TPO) is involved in autoimmune thyroid diseases and high titers of TPO autoantibodies directed to various conformational B cell epitopes are frequently present in patients' sera. Deciphering these epitopes is a difficult task, but can give insight into the structural basis of autoimmune recognition. TPO is a membrane-bound enzyme with the extracellular part organized in three protein domains, but of unknown three-dimensional structure. We previously localized a TPO B cell epitope within amino acid residues 742-848, a region encompassing the two C-terminal, extracellular domains of the protein. We found that at least one of the three tyrosine residues of the peptide 742-848 might be involved in autoantibody binding. In this study, we show by site-directed mutagenesis that the autoepitope contains tyrosine 772 located near the hinge area between the two protein domains, suggesting they are both involved in the epitope structure. The B cell epitopes of TPO are clustered in two overlapping immunodominant regions. To map the newly localized epitope with respect of these regions, competition experiments were performed using a reference panel of TPO mAb and a further mAb previously found to be specific for the TPO peptide 742-848 at variance with all the other ones. Here, we show that the tyrosine 772-bearing epitope in the peptide 742-848 maps in a region that partly overlaps the reported two immunodominant regions. These results are suggestive of a complex TPO folding that involves all the three TPO protein domains to form a highly conformational immunodominant region.
Estienne
Valérie
V
U555 INSERM/Laboratoire de Biochimie Endocrinienne et Métabolique, Faculté de Médecine, Université de la Méditerranée, 13385 Marseille Cedex 05, France.
Duthoit
Christine
C
Blanchin
Stéphanie
S
Montserret
Roland
R
Durand-Gorde
Josée-Martine
JM
Chartier
Martine
M
Baty
Daniel
D
Carayon
Pierre
P
Ruf
Jean
J
D000818:Animals; D016466:CHO Cells; D002942:Circular Dichroism; D006224:Cricetinae; D017931:DNA Primers; D018604:Epitope Mapping; D018985:Epitopes, B-Lymphocyte / Q000737:chemistry*; D006801:Humans; D016056:Immunodominant Epitopes / Q000378:metabolism; D007453:Iodide Peroxidase / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D018448:Models, Immunological; D008958:Models, Molecular; D008968:Molecular Conformation; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000032:analysis / Q000378:metabolism; D014162:Transfection; D014443:Tyrosine / Q000032:analysis* / Q000378:metabolism
D017931:DNA Primers; D018985:Epitopes, B-Lymphocyte; D016056:Immunodominant Epitopes; D011994:Recombinant Proteins; D014443:Tyrosine; D007453:Iodide Peroxidase