Purification of Nm23-H1 fusion protein expression and its function

张志敏; 杨雪琴; 许文; 戴楠; 曾林立; 李增鹏; 王东; 王阁

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Purification of Nm23-H1 fusion protein expression and its function

更新时间：2026-01-09

- Purification of Nm23-H1 fusion protein expression and its function
- Journal of Army Medical University Vol. 32, Issue 17, Pages: 1824-1827(2010)
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  CLC： R394-33;R394.3;R784
- Published：2010
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张志敏,杨雪琴,许文,戴楠,曾林立,李增鹏,王东,王阁.肿瘤转移抑制基因Nm23-H1融合蛋白的表达纯化及其功能的初步研究[J].陆军军医大学学报(原第三军医大学学报),2010,32(17):1824-1827.

　Zhang Zhimin,Yang Xueqin,Xu Wen,Dai Nan,Zeng Linli,Li Zengpeng,Wang Dong,Wang Ge.Purification of Nm23-H1 fusion protein expression and its function[J].J Amry Med Univ (J Third Mil Med Univ),2010,32(17):1824-1827.
张志敏,杨雪琴,许文,戴楠,曾林立,李增鹏,王东,王阁.肿瘤转移抑制基因Nm23-H1融合蛋白的表达纯化及其功能的初步研究[J].陆军军医大学学报(原第三军医大学学报),2010,32(17):1824-1827. DOI：

　Zhang Zhimin,Yang Xueqin,Xu Wen,Dai Nan,Zeng Linli,Li Zengpeng,Wang Dong,Wang Ge.Purification of Nm23-H1 fusion protein expression and its function[J].J Amry Med Univ (J Third Mil Med Univ),2010,32(17):1824-1827. DOI：

摘要

目的利用人Nm23-H1基因原核表达载体，在大肠杆菌中表达并纯化融合蛋白，并对蛋白其活性进行初步分析。方法将重组质粒pET28a-Nm23-H1转化入E.coli BL21(DE3)，IPTG诱导表达并鉴定。用金属螯合层析技术进行蛋白纯化，得到Nm23-H1融合蛋白，用SDS-PAGE及Western blot鉴定纯化蛋白，并用RP-HPLC法、电泳迁移率变动分析实验检测Nm23-H1融合蛋白的核苷二磷酸激酶（NDPK）活性及核酸内切酶（AP）修复活性。结果转化溶原菌后能够表达目的蛋白，蛋白表达量高，为可溶性蛋白。Ni

柱纯化后得到Nm23-H1融合蛋白经SDS-PAGE及Western blot鉴定正确。RP-HPLC法就及电泳迁移率变动分析实验证明所纯化的Nm23-H1融合蛋白具有NDPK活性，不具有AP修复活性，但可以增加APE1蛋白的AP修复活性。结论利用原核表达载体pET28a-Nm23-H1在E.coli BL21(DE3)中高效表达和纯化得到具有NDPK活性，但无AP修复活性的Nm23-H1融合蛋白，此蛋白可以增强APE1蛋白的AP修复活性，共同参与细胞的DNA损伤修复。

Abstract

Objective　To construct the prokaryotic expression vector for human Nm23-H1 genes and purify its fusion protein and study its activity.　Methods　Recombinant plasmid pET28a-Nm23-H1 was transformed into E.coli BL21 (DE3) and its expression was induced and identified with IPTG. The expressed Nm23-H1 fusion protein was purified by affinity chromatographic with Ni

column and identified by SDS-PAGE and Western blotting. Immunogenicity and activity of Nm23-H1 fusion protein were analyzed by RP-HPLC and EMSA.　Results　The prokaryotic expression vector of pET2

8a-Nm23-H1 could express the Nm23-H1 fusion protein in E.coli BL21 (DE3). The Nm23-H1 fusion protein with NDPK activity could be purified by affinity chromatographic with Ni

column. However

the AP endonuclease activity of Nm23-H1 protein was too weak to be detected.　Conclusion　Nm23-H1 fusion protein is highly expressed in E.coli BL21 (DE3) and can be purified，which is of great significance for elucidating the biologic characteristics of Nm23-H1 genes.

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