Expression And Characterization Of Recombinant Lysozyme From The Sea Cucumber Stichopus Japonicus

The study focused on fermentation and characterization of the genetically engineered bacteria E. coli Rosetta(DE3)pLysS/pET-32a(+)-SjLys to express the recombinant lysozyme from the sea cucumber Stichopus japonicus (rSjLys). The rSjLys was expressed in solube form via optimizing fermentation media and conditions. The optimal medium composition of fermenting the engineering strain was determined by orthogonal test and the product of the rSjLys was purified by Ni2+metal affinity chromatography. The production yield of the rSjLys was obtained as the range of30-40mg rSjLys in1L culture solution in shaking flasks under the fermenting conditions of induction temperature at30℃, rotational speed at140r/min, concentration of IPTG at0.3mmol/L and culturing for14h. The analysis of SDS-PAGE was used to verify the availability of the rSjLys. The results in the study proved that the optimal fermentation conditions were very helpful for over-expressing target protein.The antibacterial activity of the purified recombinant proteins of rSjLys was detected according to agarose radial diffusion assay. The results showed that the recombinant proteins of rSjLys have an inhibitive activity on Micrococcus lysodeikticus Vibrio parahaemolyticus and S.aureus. In addition, the recombinant protein rSjLys after heating treatment could improve21～52%of activities against M. lysodeikticus V. parahaemolyticus and S.aureus. These results have demonstrated that the genetically engineering strain E. coli Rosetta(DE3)pLysS/pET-32a(+)-SjLys could highly produce the recombinant protein rSjLys, with features of soluble products and potent antibacterial activity. So they will be widely used in aquaculture, livestock and poultry breeding, feed additives and food processing industry.. Therefore, the products in this study have great market potential and development value.