Expression, purification and NETs degradation activity of human DNase I

LIANG Yixuan; WU Jie

doi:10.11665/j.issn.1000-5048.20190113

Journal of China Pharmaceutical University > 2019 > 50(1): 93-99. > DOI: 10.11665/j.issn.1000-5048.20190113

LIANG Yixuan, WU Jie. Expression, purification and NETs degradation activity of human DNase I[J]. Journal of China Pharmaceutical University, 2019, 50(1): 93-99. DOI: 10.11665/j.issn.1000-5048.20190113

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Expression, purification and NETs degradation activity of human DNase I

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Abstract

In order to study the effect of human deoxyribonuclease I(DNase I)on neutrophil extracellular traps(NETs)degradation, genetic engineering bacteria E. coli Rosetta(DE3)/pET32a-His-DNase I was constructed. The fusion protein His-DNase I was induced by lactose, and purified by Ni-affinity chromatography. Mouse neutrophils were extracted and stimulated with phorbol-12-myristate-13-acetate(PMA)to form NETs. The degradation activity of the fusion protein on NETs was detected by SytoxGreen and fluorescence microscopy. The results showed that the purified His-DNase I had high nuclease activity. This study provided the research foundation for further exploration of the clinical application of DNase I.
- deoxyribonuclease I,
- neutrophil extracellular traps,
- separation and purification,
- His-tag

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