RESUMO
@#Objective To express and purify Cc PT1 protein from Aspongopus chinensis in prokaryotic cell.Methods Thesynthesized Cc PT1 gene was cloned to vector p GEX-4T-1 to construct recombinant expression plasmid p GEX-4T1-Cc PT1,which was then transformed to competent E.coli Rosetta strain and induced by IPTG.The induction temperature(20 ℃ and37 ℃),final concentration of IPTG(0.25,0.5,0.75 and 1 mmol/L)and induction time(6,8,10,12 h)were opti-mized.The obtained protein was purified by GST protein purification system,which was then analyzed by 10% SDS-PAGEand identified by Western blot.GST tags were removed by Pre Scission Protease during purification.Results The recombi-nant protein GST-Cc PT1 was expressed in the form of inclusion body with a concentration of 0.026 9 mg/ml,of which therelative molecular mass was 29 800,consistent with the expectation.The optimum induction condition was induction withIPTG of final concentration of 0.75 mol/L for 12 h at 20 ℃.The purified protein was more than 90% in purity and boundspecifically to mouse monoclonal antibody against GST.After remove of GST tags,Cc PT1 protein showed a relative molecu-lar mass of about 2 830 and the yield was 11.15%.Conclusion A.chinensis Cc PT1 protein was expressed by prokaryoticexpression system,and the purity of Cc PT1 protein was high after purification,which laid a foundation of the in-depth studyof anticancer peptides of A.chinensis.