Preparation of recombinant endolysin from Vibrio parahaemolytics-infecting bacteriophage Lys qdvp001 CHAP domain portein in Escherichia coli and preliminary optimization of inducing conditions

Objective:In order aimed to use E.Coli BL21(DE3)to over express Lys qdvp001 CHAP recombinant and produce soluble pET30a-CHAP protein using renaturation. Methods:PET30a-CHAP expression vector was constructed,and to express the recombinant protein,IPTG was added. Then detect the protein of supernatant in order to know the form of protein expressing. After that,the optimal expression condition was studyed. PET30a-CHAP inclusion body protein was washed once by using detergent to get rid of protein impurity and then dissolved by Uera. After purification by Ni²⁺ Sepharose^TM 6 Fast Flow affinity chromatography column,the purified protein was dialyzed by renaturation accelerator(EDTA,oxidized glutathione and glutathione asxpression)to obtain soluble pET30a-CHAP protein. Finally,detect the activity of the protein. Results:The results shows recombinant protein is in the inclusion body.The optimal expression condition was Incubation with IPTG(0.5 mmol/L)at 16 ℃ for 7 h. And the protein can slower the growth of Vibrio parahaemolyticus. Conclusion:This research would provide a method for the proparation of endolysin to prevent the infection of Vibrio parahaemolyticus and other gram negative bacteria.