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. 2005 Mar;21(2):167-70.

[Improvement of refolding efficiency of HLA-A2-peptide complex in vitro by preformed disulfide bonds in heavy chain]

[Article in Chinese]
Shi-jiang Fei  1 , Xiong-wen Wu, Xiu-fang Wong, Lei Cai, Zhi-hui Liang, Jun-yan Han, Fei-li Gong
Affiliations
  • PMID: 15766400

[Improvement of refolding efficiency of HLA-A2-peptide complex in vitro by preformed disulfide bonds in heavy chain]

[Article in Chinese]
Shi-jiang Fei et al. Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2005 Mar.

Abstract

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 (NH4)2SO4 precipitation were applied to purify the HC. Then the purified HC, beta2m 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 beta2m antibody.

Results: The refolded products consisted of HLA-A2-peptide complex, beta2m, 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.

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