Use of domain-swapped molecules for conformational epitope mapping of desmoglein 3 in pemphigus vulgaris.
Pemphigus vulgaris is an autoimmune blistering disease caused by autoantibodies against desmoglein 3, a member of the desmosomal cadherin family. These autoantibodies recognize conformation-dependent epitopes on desmoglein 3. In this study we attempted to map the conformational epitopes of desmoglein 3 in pemphigus vulgaris using recombinant desmoglein 3 produced by the baculovirus expression system. We developed a series of domain-swapped molecules between desmoglein 3 and desmoglein 1, which have similar structures but distinct epitopes. These were developed by substituting deleted segmental regions of desmoglein 3 by the corresponding desmoglein 1. Thus domain-swapped molecules containing desmoglein 3 residues 1-403, 1-161, 163-566, and 405-566 were constructed and used as competitors for competition enzyme-linked immunosorbent assay against the entire extracellular domain of desmoglein 3 with 25 pemphigus vulgaris sera. Considering more than 50% absorption as significant, residues 1-403 and 1-161 showed significant absorption in 24 out of 25 (96%) and 18 out of 25 (72%) pemphigus vulgaris sera, respectively, whereas only one serum and no sera showed significant absorption by residues 163-566 and 405-566, respectively. Furthermore, no apparent change in their major epitopes was seen during the time course in four pemphigus vulgaris cases tested. These findings indicate that the domain-swapping approach is useful for conformational epitope mapping in pemphigus and that amino-terminal residues 1-161, which are considered to include a region essential for cell-cell adhesion in cadherins, contain the critical residues of the conformational epitope of desmoglein 3 recognized by the autoantibodies in pemphigus vulgaris sera.
Futei
Y
Y
Department of Dermatology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan.
Amagai
M
M
Sekiguchi
M
M
Nishifuji
K
K
Fujii
Y
Y
Nishikawa
T
T
D001324:Autoantigens / Q000276:immunology; D015820:Cadherins / Q000276:immunology*; D051184:Desmoglein 3; D018604:Epitope Mapping / Q000379:methods*; D006801:Humans; D010392:Pemphigus / Q000276:immunology*; D017434:Protein Structure, Tertiary / Q000502:physiology; D013997:Time Factors
D001324:Autoantigens; D015820:Cadherins; C071894:DSG3 protein, human; D051184:Desmoglein 3