MIP-3α-SA双功能融合蛋白的制备及其生物学功能鉴定

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Abstract Objective: To prepare the streptavidin-tag MIP-3α fusion protein MIP-3α-SA, and to study its biological function and activity. Methods: The recombinant expression vector of MIP-3α-SA-pET21 was constructed and induced to express MIP-3α-SA fusion protein in Escherichia coli. MIP-3α-SA fusion protein was purified by the Ni-NTA affinity chromatography and refolded by gradient dialysis of urea. The fusion protein was identified by amide gel electrophoresis (SDS-PAGE), Western blot and silver ammonia staining. In vitro lymphocyte chemotaxis assays was used to detect the chemotactic activity of the fusion protein, and flow cytometry (FCM) analysis was performed to detect the modification efficiency of the fusion protein on biotin RM-1 cell surface anchoring. Results: The MIP-3α-SA fusion protein expression was induced with high efficiency in E. coli Rosetta (DE3), which accounted for 30% of the total bacterial protein. The purity of the fusion protein was 95% after purification by nickel column affinity chromatography. After renaturation by urea gradient, the fusion protein was verified to have dual activity, namely: MIP-3α-SA stimulated human lymphocyte chemotactic activity has a dose dependent manner and SA-mediated efficient binding to the surface has biotinylated RM-1 cell function (modification of the surface anchoring efficiency of more than 95%). Conclusion: The MIP-3α-SA bifunctional fusion protein has dual activity, which lays the foundation for the research and development of MIP-3α surface-modified tumor cell vaccine.

Key words： macrophage inflammatory protein-3α streptavidin fusion protein anchoring modification

Received: 11 October 2018

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	SONG Zhichun
	DAI Shu
	WANG Peng
	GAO Jimin.

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https://xb.wmu.edu.cn/EN/ OR https://xb.wmu.edu.cn/EN/Y2019/V49/I5/360

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