Auto-induction for high level production of biologically active reteplase in Escherichia coli

Publication date: November 2018 Source:Protein Expression and Purification, Volume 151 Author(s): Mehrnoosh Fathi-Roudsari, Nader Maghsoudi, Amirhossein Maghsoudi, Sepideh Niazi, Morvarid Soleiman Reteplase is a third generation tissue plasminogen activator (tPA) with a modified structure and prolonged half-life in comparison to native tPA. As a non-glycosylated protein, reteplase is expressed in Escherichia coli. Due to presence of several disulfide bonds, high level production of reteplase is complicated and needs extra steps for conversion to biologically active form. Auto-induction represents a method for high-yield growth of bacterial cells and higher expression of recombinant proteins. Here we have tried to optimize the auto-induction procedure for soluble and active expression of reteplase in E. coli. Results showed that using auto-induction strategy at 37 °C, Rosetta-gami (DE3) had the highest level of active and soluble reteplase production in comparison to E. coli strains BL21 (DE3), and Shuffel T7. Temperature dominantly affected the level of active reteplase production. Decreasing the temperature to 25 and 18 °C increased the level of active reteplase by 20 and 60%, respectively. The composition of auto-induction medium also dramatically changed the active production of reteplase in cytoplasm. Using higher enriched auto-induction medium, super broth base including trace elements, significantly increased biologically active reteplase by 30%. It is demo...
Source: Protein Expression and Purification - Category: Biochemistry Source Type: research