Heterologous expression,refolding,and characterization of a calcium-independent phospholipase A1 from Streptomyces albidoflavus  

作　　者：Shi Cheng Zitao Guo Chaojuan Liang Yi Shi Peng Geng Yu Xin Zhenghua Gu Liang Zhang 

机构地区：[1]National Engineering Laboratory for Cereal Fermentation Technology,Jiangnan University,Wuxi 214122,Jiangsu,People’s Republic of China

出　　处：《Systems Microbiology and Biomanufacturing》2022年第3期487-497,共11页系统微生物学与生物制造（英文）

基　　金：supported by the National Key Research&Developmental Program of China(2021YFC2100303,2018YFA0900300).

摘　　要：Phospholipase A1(PLA_(1))is a member of the hydrolase family with applications in various fields,especially in the food industry.A calcium-independent PLA_(1) from Streptomyces albidoflavus was expressed in E.coli BL21(DE3)in this study.The results indicated that the soluble expression of PLA_(1) was at low level,which was possibly due to the toxicity of PLA_(1) to the host.In contrast,the expression of the enzyme as inclusion bodies exhibited a high-level expression and 0.3 mg inclusion bodies protein could be derived from 1 mL culture medium.Furthermore,the renaturation of PLA_(1) was achieved through a direct dilution method,yielding 29.6 U/mL PLA_(1) activity after 16 h of renaturation at 4℃.For improving the efficiency of the dilution refolding process,a continuous refolding strategy was established,and 155 U/mL PLA_(1) activity was derived from the continuous refolding process.With soybean lecithin as the substrate,the specific activity of purified renatured PLA_(1) was 1380 U/mg and the optimal temperature and pH was found to be 60℃ and 6.5.In addition,the renatured PLA_(1) was observed with better activity towards phosphatidyl inositol,whilst lipase activity was detected when the catalyzing temperature was below 55℃.Overall,this study provides a possible solution to obtain calcium-independent PLA_(1) with high yield by heterologous expression in E.coli and hence to promote its further application in the field of food industry.

关 键 词：Calcium-independent phospholipase A1 Heterologous expression Continuous refolding CHARACTERIZATION Escherichia coli 

分 类 号：Q55[生物学—生物化学]