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预先形成的重链二硫键有助于HLA-A2-抗原肽复合物体外折叠 被引量：1

Improvement of refolding efficiency of HLA-A2-peptide complex in vitro by preformed disulfide bonds in heavy chain
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摘要 　目的: 提高可溶性HLA -A2 肽复合物体外折叠效率。方法: 在变性非还原条件下提取原核表达的HLA重链(HC)。通过离子交换及硫酸铵沉淀初步纯化后, 在β2微球蛋白(β2m)及特异性抗原肽 (Try369-377 )存在的情况下, 于pH6. 6的折叠缓冲体系中稀释复性。利用Westernblot及ELISA检测折叠产物。结果: 折叠复合物中主要含有HLA-A2 抗原肽复合物和β2m, 较少含有HC聚合体; 折叠效率较传统方法提高 2. 5倍。结论: 通过与传统方法作比较, 证实此法折叠效率比传统方法高, 为进一步HLA 肽四聚体及人工抗原提呈细胞的制备可提供足够可溶性的HLA -A2 肽单体。 AIM: To improve the refolding efficiency of soluble HLA-A2-peptide complex in vitro. METHODS: The heavy chain (HC) of MHC class I was extracted from bacteria under denaturing and non-reducing conditions. Anion-exchange and (NH 4) 2SO 4 precipitation were applied to purify the HC. Then the purified HC, β 2m and an antigenic peptide (N-YMDGTMSQV-COOH of Try 369-377) were refolded to form an HLA-A2-peptide complex by dilution method in the buffer of pH 6.6. The refolded products were detected by Western blot and ELISA with W6/32 and anti-human β 2m antibody. RESULTS: The refolded products consisted of HLA-A2-peptide complex, β 2m, and a little amount of HC polymer. The refolding efficiency was 2.5 fold higher than that of the conventional method. CONCLUSION: This study confirmed that the refolding efficiency of the method reported in this paper is higher as compared with the conventional method, which is of importance to the preparation of HLA-peptide tetramers and artificial antigen presenting cells.
作者 费世江 吴雄文 翁秀芳 蔡蕾 梁智辉 韩军艳 龚非力
机构地区 华中科技大学同济医学院免疫学系
出处 《细胞与分子免疫学杂志》 CAS CSCD 北大核心 2005年第2期167-170,共4页 Chinese Journal of Cellular and Molecular Immunology
基金 国家自然科学基金资助项目 (No. 30271201 ) 国家重点基础研究发展规划(973)资助项目(No.2001CB510008)
关键词 HLA-A2-抗原肽复合物 体外折叠 二硫键形成 HLA-A2-peptide complex in vitro folding disulfide bond formation
分类号 R392.12 [医药卫生—免疫学]
  • 相关文献

参考文献12

  • 1翁秀芳,陈浩,吴雄文,梁智辉,韩军艳,黄亚非,龚非力.可溶性HLA-A2-肽复合物的体外折叠与生物素化[J].细胞与分子免疫学杂志,2004,20(3):265-268. 被引量：10
  • 2Garboczi DN, Hung DT, Willy DC, et al. HLA-A2-peptide complexes:refolding and crystallization of molecules expressed in Escherichia coli and complexed with single antigenic peptides[J]. Proc Natl Acad Sci USA, 1992, 89: 3429-3433.
  • 3Altaman JD, Moss PH, Goulder PR, et al. Phenotypic analysis of antigen-specific T lymphocytes[J]. Science, 1996, 274: 94-96.
  • 4Constantin CM, Bonney EE, Altman JD, et al. Major histocompatibility complex (MHC) tetramer technology: an evaluation[J]. Biol Res Nurs, 2002, 4(2): 115-127.
  • 5Parker KC, Silver ML, Wiley DC, et al. An HLA-A2/β2m/peptide complex assembled from subunits expressed separately in E.coli[J]. Mol immunol, 1992, 29(3): 371-378.
  • 6Rudolph R, Lilie H. In vitro folding of inclusion body proteins[J]. FASEB J, 1996, 10: 49-56.
  • 7Pedersen LQ, Nissen MH, Hansen NJ, et al. Efficient assembly of recombinant major histocompatibility complex class I molecules with preformed disulfide bonds[J]. Eur J Immunol, 2001, 31: 2986-2996.
  • 8Maziarz RT, Fraser J, Srominger JL, et al. The human HLA-specific monoclonal antibody W6/32 recognizes a discontinuous epitope within the alpha 2 domain of murine H-2Db[J]. Immunogenetics, 1986, 24(3): 206-208.
  • 9Ferre H, Ruffet E, Blicher T, et al. Purification of correctly oxidized MHC class I heavy-chain molecules under denaturing conditions: A novel strategy exploiting disulfide assisted protein folding[J]. Protein Science, 2003, 12: 551-559.
  • 10Sylvester-Hvid C, Kristensen NS, Blicher T, et al. Establishment of a quantitative ELISA capable of determining peptide-MHC class I interaction[J]. Tissue Antigen, 2002, 59: 251-258.

二级参考文献9

  • 1Garboczi DN, Hung DT, Wiley DC, et al. HLA-A2-peptide complexes: refolding and crystallization of molecules expressed in Escherichia coli and complexed with single antigenic peptides[J]. Proc Natl Acad Sci USA, 1992, 89(8): 3429-3433.
  • 2Altman J, PAH Moss P, Goulder D, et al. Phenotypic analysis of antigen-specific T lymphocytes[J]. Science, 1996, 274(5284): 94-96.
  • 3McMichael AJ, Callaghan CA. A new look at T cells[J]. J Exp Med, 1998, 187(9): 1367-1371.
  • 4Tham EL, Jensen PL, Mescher MF. Activation of antigen-specific T cells by artificial cell construcs having immobilized multimeric peptide-class I complexes and recombinant B7-Fc protein[J]. J Immunol Methods, 2001, 249(1-2): 111-119.
  • 5Maziarz RT, Fraser J, Srominger JL, et al. The human HLA-specific monoclonal antibody W6/32 recognizes a discontinuous epitope within the alpha 2 domain of murine H-2Db[J]. Immunogenetics, 1986, 24: 206-208.
  • 6Ocallaghan CA, Byford MF, Wyer JR, et al. BirA enzyme: production and application in the study of membrane receptor-ligand interactions by site-specific biotinylation[J]. Anall Biochem, 1999, 266(1): 9-15.
  • 7Parker KC, Silver ML, Wiley DC. An HLA-A2/β2m/peptide complex assembled from subunits expressed separately in E.coli[J]. Mol Immunol, 1992, 29(3): 371-378.
  • 8吴涛,兰炯采,张泓,王从容,毛高才.ELISA法检测可溶性人类白细胞抗原-Ⅰ及广东人参考值的测定[J].第一军医大学学报,2002,22(10):931-933. 被引量：2
  • 9宁双飞,贾卫,张新海,杨琨,李琦,刘雪松,金伯泉.检测可溶型CD226双mAb夹心ELISA的建立及初步应用[J].细胞与分子免疫学杂志,2003,19(2):192-194. 被引量：3

共引文献9

同被引文献8

  • 1Watanabe N,Gavrieli M,Sedy JR,et al.BTLA is a lymphocyte inhibitory receptor with similarities to CTLA-4 and PD-1[J].Nat Immunol,2003,4(7):670-679.
  • 2Han P,Goularte OD,Rufner K,et al.An inhibitory Ig superfamily protein expressed by lymphocytes and APCs is also an early marker of thymocyte positive selection[J].J Immunol,2004,172(10):5931-5939.
  • 3Sedy JR,Gavrieli M,Potter KG,et al.B and T lymphocyte attenuator regulates T cell activation through interaction with herpesvirus entry mediator[J].Nat Immunol,2005,6(1):90-98.
  • 4Gonzalez LC,Loyet KM,Calemine-Fenaux J,et al.A coreceptor interaction between the CD28 and TNF receptor family members B and T lymphocyte attenuator and herpesvirus entry mediator[J].Proc Natl Acad Sci USA,2005,102(4):1116-1121.
  • 5Shen Z,Reznikoff G,Dranoff G,et al.Cloned dendritic cells can present exogenous antigens on both MHC class I and class II molecules[J].J Immunol,1997,158(6):2723-2730.
  • 6Wang Y,Subudhi SK,Anders RA,et al.The role of herpesvirus entry mediator as a negative regulator of T cell-mediated responses[J].J Clin Invest,2005,115(3):711-717.
  • 7Compaan DM,Gonzalez LC,Tom I,et al.Attenuating lymphocyte activity:the crystal structure of the BTLA-HVEM complex[J].J Biol Chem,2005,280(47):39553-39561.
  • 8吴东,沈锋,娄永华,焦炳华,吴孟超.人LIGHT基因的克隆及其重组腺病毒载体的构建[J].细胞与分子免疫学杂志,2002,18(2):121-124. 被引量：8

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细胞与分子免疫学杂志
2005年 第2期

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