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Expression of human myeloid differentiation protein-2 in E.coli
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- Author:
- No author available
- Journal Title:
- JOURNAL OF THE FOURTH MILITARY MEDICAL UNIVERSITY
- Issue:
- 14
- DOI:
- 10.3321/j.issn:1000-2790.2004.14.003
- Key Word:
- 髓样分化蛋白-2;基因表达;重组融合蛋白质类;谷胱甘肽转移酶
Abstract： 目的:在大肠杆菌DH-5α中表达人类髓样分化蛋白-2(human myeloid differentiaton protein-2,hMD-2)与谷胱甘肽巯基转移酶(glutathione-s-transferase,GST)的融合蛋白(GST/hMD-2),并对融合蛋白进行免疫学鉴定及初步的变性复性处理.方法:建立GST/hMD-2表达菌,在不同温度下以不同浓度IPTG诱导不同时间后采样,用SDS-PAGE分析各种组合条件下融合蛋白的表达量和溶解性;用Western-Blot鉴定融合蛋白免疫性;用尿素变性和透析法对融合蛋白进行初步纯化.结果:在37℃经0.4 mmol/L IPTG诱导4 h后,GST/hMD-2可获最高表达量(约占菌体总蛋白的20%),未见可溶性表达;Western-Blot证实GST/hMD-2能与抗MD-2单克隆抗体特异性结合;GST/hMD-2溶于8 mol/L尿素,梯度透析后纯度提高至40%.结论:hMD-2可在大肠杆菌DH-5α中与GST融合表达.
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