Expression of Pleurocidin from winter flounder in Escherichia coli and optimization of culture conditions / 生物工程学报

To express Pleurocidin in Escherichia coli and to enhance the secretory efficiency of the fusion protein, the gene encoding Pleurocidin was ligated with Cherry DNA sequence via blunt-end ligation. Then this fusion gene was cloned into pET22b (+) vector and the recombinant plasmid was transformed into E. coli BL21 (DE3). Lactose was used to induce expression of fusion protein. The recombinant plasmid pET22b (+) -CP was successfully constructed and high-level expression of fusion protein was induced with lactose. Statistics showed that addition of glycine after 16 h of induction significantly enhanced the secretory efficiency of the fusion protein. After hydrolysis of the fusion protein by diluted hydrochloric acid and some further purification steps, r-Pleurocidin was obtained with antibacterial activity against E. coli DH5α and Bacillus subtilis BS168. In conclusion, the fusion protein was expressed in E. coli and biologically active r-Pleurocidin was obtained after hydrochloric acid cleavage and purification.

Research and development of enzymes used in feed / 生物工程学报

Enzymes can degrade the anti-nutrient factors in feedstuff, increase nutrient digestibility, and reduce pollution to environment, and have been widely supplemented in animal feedstuff. However, the use of enzymes is limited because of their undesirable properties, such as thermoliability and susceptibility against protease digestions. And its commercialization is also limited by low production efficiency and high cost. Therefore, the focuses for future enzyme development will be: (1) to obtain novel enzymes with better properties by high-throughput screening of enzyme encoding genes, especially those from extreme and special environments; (2) to improve enzyme properties using directed mutagenesis and protein engineering methods; (3) to achieve high-level fermentation of enzymes by heterogonous expression and optimization of codons, vectors and fermentation conditions; (4) to determine the effect of enzymes to animals and utilize enzymes efficiently.