Expression and Purification of Neurotoxin-Associated Protein HA-33/A from Clostridium botulinum and Evaluation of Its Antigenicity

Authors
Ali Sayadmanesh
Firouz Ebrahimi
Abbas Hajizade
Mosayeb Rostamian
Hani Keshavarz
faculty member of IHU
10.6091/ibj.1216.2013
Abstract
Background: Botulinum neurotoxin (BoNT) complexes consist of neurotoxin and neurotoxin-associated proteins. Hemagglutinin-33 (HA-33) is a member of BoNT type A (BoNT/A) complex. Considering the protective role of HA-33 in preservation of BoNT/A in gastrointestinal harsh conditions and also its adjuvant role, recombinant production of this protein is favorable. Thus in this study, HA-33 was expressed and purified, and subsequently its antigenicity in mice was studied. Methods: Initially, ha-33 gene sequence of Clostridium botulinum serotype A was adopted from GenBank. The gene sequence was optimized and synthesized in pET28a (+) vector. E. coli BL21 (DE3) strain was transformed by the recombinant vector and the expression of HA-33 was optimized at 37°C and 5 h induction time. Results: The recombinant protein was purified by nickel nitrilotriacetic acid agarose affinity chromatography and confirmed by immunoblotting. Enzyme Linked Immunoassay showed a high titer antibody production in mice. Conclusion: The results indicated a highly expressed and purified recombinant protein, which is able to evoke high antibody titers in mice.
Keywords
Botulinum neurotoxin
Expression
Purification