Production of the entire extracellular domain of BP180 (type XVII collagen) by baculovirus expression.
Bullous pemphigoid (BP) is an acquired autoimmune skin disease, and its target antigens are a 230 kDa plaque protein (BP230) and a 180 kDa transmembrane protein with interrupted collagenous domains (BP180, type XVII collagen), which localize at the hemidesmosome. In this study we have attempted to express the entire extracellular domain of BP180 (rBP180EC) as a secreted protein by baculovirus expression. Seventy out of 83 BP sera (84.4%) showed positive reactivity against rBP180EC by immunoblot analysis, and 56 out of 83 BP sera (67.5%) were positive against rBP180EC by ELISA. These figures were comparable with those when a bacterial recombinant protein encoding the NC16a domain of BP180 (rNC16a) was used as an antigen source. Reactivity of BP sera against rBP180EC by ELISA was completely abolished or significantly reduced by immunocompetition with rNC16a in 11 out of 14 BP sera tested, while the reactivity was not altered in the rest of the three sera. These findings indicate that the NC16a domain represents the major epitopes on the extracellular domain of BP180, although there are some other minor epitopes outside of NC16a which are uniquely expressed by rBP180EC. rBP180EC will be useful to develop a diagnostic tool for BP as well as to dissect a molecular role for BP180 in interactions of keratinocytes with epidermal basement membrane.
Hata
Y
Y
Department of Dermatology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan. hata@mc.med.keio.ac.jp
Fujii
Y
Y
Tsunoda
K
K
Amagai
M
M
D001324:Autoantigens / Q000235:genetics*; D016367:Baculoviridae; D002352:Carrier Proteins; D003094:Collagen / Q000235:genetics*; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D004926:Escherichia coli; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D010391:Pemphigoid, Bullous / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D000097805:Collagen Type XVII
D001324:Autoantigens; D002352:Carrier Proteins; D003094:Collagen; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D011994:Recombinant Proteins; D000097805:Collagen Type XVII; C486268:DST protein, human