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蓝氏贾第鞭毛虫沉默信息调节因子2重组蛋白复性方法比较
  • ISSN号:1673-5234
  • 期刊名称:《中国病原生物学杂志》
  • 时间:0
  • 分类:R382.2[医药卫生—医学寄生虫学;医药卫生—基础医学]
  • 作者机构:[1]大连大学医学研究中心分子生物学实验室,辽宁大连116622
  • 相关基金:国家自然科学基金项目(No.30872207,No.30901253).
中文摘要:

目的采用3种不同复性方法获得具有生物活性的重组蛋白蓝氏贾第鞭毛虫(Giardia lamblia,G1)沉默信息调节因子2(GISir2),并比较3种方法的复性效率,为研究其活性和生物学功能奠定基础。方法表达并纯化G1Sir2的包涵体,分别采用稀释复性、透析复性和柱上复性等3种方法对G1Sir2包涵体进行复性,并通过检测复性蛋白的活性,比较3种复性方法的特点。结果蛋白活性回收以柱上复性较高,为1712Flu,透析复性和稀释复性法分别为758F1u和206Flu。结论3种复性方法均能获得有活性的融合蛋白G1Sir2,其中以柱上复性方法处理的蛋白活性回收值最高。

英文摘要:

Objective To compare the refolding efficiency of recombinant silent information regulator 2 (G1Sir2) protein from Giardia larnblia by three different methods of refolding and to determine the most effective way to obtain biological- ly active G1Sir2 protein. Methods Recombinant G1Sir2 was expressed in Escherichia. coZi. The inclusion bodies were collected and dissolved with 8 mol/L urea. The supernatant was used in Ni~+ affinity purification or directly in on-column refolding. The purified G1Sir2 was refolded by two different methods, dialysis refolding or dilution refolding. G1Sir2 pro- teins refolded by three different methods were analyzed with 12~ SD~PAGE and deacetylase activity was determined with a Histone Deacetylase Assay Kit (Fluorometric) in vitro. The rate of G1Sir2 refolding was measured and rate of re- covery was calculated. Results G1Sir2 protein refolded by on-column-refolding had the highest deacetylase-specifie ac- tivity, followed by the protein refolded by dilution and dialysis. The highest rate of G1Sir2 activity was achieved by oncolumn refolding, followed by dialysis and dilution; the level of activity was 1 712, 758, and 206 Flu, respectively. Conclusion Active GISir2 protein can be obtained by all three methods of refolding though on-column refolding is preferable to the other two.

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期刊信息
  • 《中国病原生物学杂志》
  • 中国科技核心期刊
  • 主管单位:中华人民共和国卫生和计划生育委员会
  • 主办单位:中华预防医学会 卫生部疾病控制司 山东省寄生虫病防治研究所
  • 主编:庄辉
  • 地址:山东省济宁市太白楼中路11号
  • 邮编:272033
  • 邮箱:www.cjpb.org
  • 电话:0537-2342934
  • 国际标准刊号:ISSN:1673-5234
  • 国内统一刊号:ISSN:11-5457/R
  • 邮发代号:24-81
  • 获奖情况:
  • 中华预防医学会、系列杂志优秀期刊,山东省优秀期刊
  • 国内外数据库收录:
  • 英国农业与生物科学研究中心文摘,中国中国科技核心期刊,中国北大核心期刊(2011版),中国北大核心期刊(2014版)
  • 被引量:8395