Heterologous expression of recombinant nattokinase in Escherichia coli BL21(DE3) and media optimization for overproduction of nattokinase using RSM

In this study, we cloned the nattokinase gene from the Bacillus subtilis strain into the pET32a vector and expressed the protein in the E. coli BL21(DE3) strain. The active recombinant nattokinase was purified using Ni-NTA affinity chromatography and then evaluated for fibrinolytic and blood clot lysis activity. Physiological parameters for optimizing protein production at optimal pH, temperature, IPTG concentration, and incubation time were investigated. A statistical technique was used to optimize media components for nattokinase overproduction, and Central Composite Design-Response Surface Methodology-based optimization was used to select significant components for protein production. The optimized media produced 1805.50 mg/L of expressed nattokinase and 42.80 gm/L of bacterial mass. The fibrinolytic activity obtained from refolded native protein was 58FU/mg, which was five times higher than the available orokinase drug (11FU/mg). The efficiency with which a statistical technique for media optimization was implemented improved recombinant nattokinase production and provides new information for scale - up nattokinase toward industrial applications.PMID:36379347 | DOI:10.1016/j.pep.2022.106198
Source: Atherosclerosis - Category: Cardiology Authors: Source Type: research