Generation of Monoclonal Antibodies Against Escherichia coli DsbA

摘要

Disulfide oxidoreductase A (also known as disulfide bond formation protein A, or DsbA) is produced in Escherichiancoli (E. coli) periplasm, which plays an important role in correct formulation of disulfide bonds duringnprotein exportation in vivo. DsbA prokaryotic expression vectors are designed for periplasmic co-expressionnof recombinant proteins with an improving secretion. In the present study, the first domain of humannCD226 (CD226D1) was expressed as a His-tagged fusion protein with DsbA and purified by Ni-NTA resin.nThree monoclonal antibodies (MAbs) against DsbA were raised by using the recombinant fusion protein asnimmunogen. We have demonstrated that one of them can detect DsbA via Western blotting, in addition to itsnability to immunoprecipitate DsbA fusion protein. Furthermore, anti-DsbA MAb can be employed to preparenantibody-coupled affinity column for purification of DsbA fusion protein from E. coli lysate.