霍乱弧菌DncV蛋白的原核表达及活性分析

doi:10.11843/j.issn.0366-6964.2015.05.026

Abstract

Abstract:

DncV（VC0179） is the member of a new family of di-nucleotide cyclases in Vibrio cholera and it can catalyzes the synthesis of c-di-AMP，c-di-GMP and c-GAMP.c-di-GMP serves as the second messenger molecule to activate the innate immunity through STING signaling pathway.To generate c-di-GMP in vitro and further investigate its functions，we constructed the propionate-inducible VC0179 prokaryotic expression plasmid fused with cherry-Tag.The recombinant plasmid was transformed into E.coil BL21（DE3） and induced by propionate.The expression of recombinant protein was observed by red color.SDS-PAGE analysis was used to identify the soluble expression of recombinant protein after ultrasonic decomposition.Moreover，c-di-GMP was enzymatically synthesized in vitro with the recombinant protein purified by Ni-NTA purification system and detected by high performance liquid chromatography.We successfully constructed the propionate-inducible VC0179 prokaryotic expression plasmid and acquired the high quality fusion protein，which could catalyze the synthesis of c-di-GMP in vitro by a one-step process.These results suggested that we established a method to generate enzymatically active VC0179 recombinant protein.The synthesis of c-di-GMP in vitro could facilitate the functional study and high volume production of c-di-GMP.

CLC Number:

S852.61

HAN Ying-qian，KONG Jiang-nan，WANG Jiang，ZHANG Chao，YANG Guo-yu. Prokaryotic Expression and Activity Analysis of DncV from Vibrio Cholera[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2015, 46(5): 868-872.

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Prokaryotic Expression and Activity Analysis of DncV from Vibrio Cholera

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