Expression and Purification of vMIP-Ⅱ N-Terminal GST Fusion Protein and Identification of Its Bioactivity

doi:Q816

China Biotechnology

2010, Vol. 30

Issue (09): 80-86 DOI: Q816

Expression and Purification of vMIP-Ⅱ N-Terminal GST Fusion Protein and Identification of Its Bioactivity

YANG Qing-ling¹,YANG Zhi-feng²,GAO Yan-jun²,CHEN Chang-jie¹

1.Department of Biochemistry & Molecular Biology,Beng Bu Medical College,Bengbu 233030,China
2.Clinical Testing and Diagnose Experimental Center of Bangbu Medical College, Bengbu 233000,China

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Abstract

The viral macrophage inflammatory protein-II (vMIP-II) is encoded by Kaposi's sarcoma-associated herpesvirus and the residues 1~21 of the N-terminal of vMIPII(NT21MP) was shown to strongly and selectively bind CXCR4 to inhibit the chemotaxis of SK-BR-3 cells.The N-terminal of vMIP-ⅡcDNA was derived by chemical sythesis method. DNA fragment encoding the NT21MP peptide was inserted into pGEX-KG vector and the recombinant plasmid was expressed in E.coli BL21(DE3). NT21MP peptide, which was recombinated with a glutathione S-transferase (GST) at the N-terminal, was expressed in E.coli cells.SDS-PAGE and Western Blot analysis demonstrated that the recombinant protein existed mainly in the supernatant of E.coli lysates. The supernatant was further purified by affinity chromatography, ultrafiltration and fast protein liquid chromatography. The bioactivity of GSTNT21MP was determined by Transwell chemotaxis on CXCR4-expressing breast cancer cell line SK-BR-3 in vitro. The results indicate that The chemotaxis of SK-BR-3 cells toward stromal cell-derived factor-1(SDF-1) was reduced by GST-NT21MP and may serve as a potential target drug for the treatment of breast cancer metastasis.

Key words： vMIP-Ⅱ CXCR4 SDF-1Purification

Received: 31 May 2010 Published: 25 August 2010

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	YANG Qing-Ling
	YANG Zhi-Feng
	GAO Yan-Jun
	CHEN Chang-Jie

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YANG Qing-Ling, YANG Zhi-Feng, GAO Yan-Jun, CHEN Chang-Jie. Expression and Purification of vMIP-Ⅱ N-Terminal GST Fusion Protein and Identification of Its Bioactivity. China Biotechnology, 2010, 30(09): 80-86.

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https://manu60.magtech.com.cn/biotech/Q816 OR https://manu60.magtech.com.cn/biotech/Y2010/V30/I09/80

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