Prokaryotic Expression,Purification and Pharmacodynamics Research of the Recombinant Protein Containing Cell-penetrating TAT Peptide and nNOS_(1-133)

Objective: The expression and purification of the recombinant protein TAT-nNOS1-133 and its effects were investigated. Methods: The prokaryotic expression vector pET28a-TAT-nNOS1-133 was constructed and transformed into the expression strain of E.coli BL21(DE3). Subsequently, the transformed bacteria were induced by lactose for expression of the recombinant protein. The recombinant protein accumulated mainly in inclusion body was purified by the sequential application of washing, renaturation, ion exchange and ultrafiltration. The transduction of the fusion protein labeled with FITC(FITC-TAT-nNOS1-133 and FITC-BSA) in cortical neurons in vitro and its neuroprotective effects were also investigated. Results: SDS-PAGE revealed that tthe recombinant protein TAT-nNOS1-133 was of high purity. TAT-nNOS1-133 can get into neurons and protect neurons against neurotoxicity. Conclusion: TAT-nNOS1-133 can be successfully expressed in prokaryotic expression system and it protects neurons in vitro.