Molecular cloning and protein expression of EC1-2 and EC3-4 epitopes of pemphigus vulgaris antigen

Objective To clone and express EC1-2 and EC3-4 epitopes of PVA ( pemphigus vulgaris antigen, desmoglein-3) in order to diagnose pemphigus and study th e relationship between epitopes of PVA and anti-PVA antibody. Methods RNA was extracted from keratinocytes and the cDNA of epitopes EC1-2 and EC3-4 was synthesized by reverse transcription. Amplified genes of EC1 -2 and EC3-4 were inserted into the expression plasmid, PGEX-4T-1, and tran sformed into E. coli BL21 by electric transduction. Recombinant fusion prot eins of EC1-2 and EC3-4 epitopes were expressed by IPTG induction. These pr oteins were separated on SDS-PAGE gels and electroblotted to nitrocellulose to detect the anti-PVA antibody. Results The sequences of cloned EC1-2 and EC3-4 genes were identical to the sequence registered in PC/GENE. Expressed recombinant proteins reacted onl y to sera from patients with pemphigus vulgaris, not to sera from patients with bullous pemphigoid, systemic lupus erythematosus or normal persons. Conclusions These recombinant proteins are very specific in antigenicity. T his may provide a new method for the diagnosis of pemphigus vulgaris (PV) o r the differential diagnosis of other bullous cutaneous diseases via patien t sera. It is also helpful in understanding the relationship between adhesi on molecules and the pathogenic mechanism of pemphigus vulgaris.