腾冲嗜酸热两面菌S5分子伴侣β亚基的表达、纯化和活性的初步分析  

The Expression,Purification of Chaperonin β Subunit from the Thermoacidophilic Archaeon,Acidianus tengchongensis and its Activity Analysis

作　　者：马晴[1] 张渝英[2] 

机构地区：[1]北京师范大学生命科学学院,北京100875 [2]中国科学院微生物研究所,北京100080

出　　处：《微生物学通报》2007年第1期28-31,共4页Microbiology China

基　　金：国家自然科学基金(No.3997003)

摘　　要：用NdeI和BamHI酶切回收腾冲嗜酸热两面菌S5的分子伴侣β亚基基因片段插入pET-23b的相应位置,并分别在BL21(DE3)和Rosetta-gami^(TM)B(DE3)pLysS中表达。表达的β亚基以可溶的形式存在。β亚基在Rosetta-gami^(TM)B(DE3)pLysS中表达较高,其占菌体总蛋白的16．2%,且以单体和聚体形式同时存在。表达的菌体经超声破碎、70℃热处理后,上清中β亚基蛋白含量达到30%,再经(NH_4)_2SO_4沉淀、Bio-Gel A-1．5m和DEAE-Sepharose CL-6B柱层析,得到在SDS-PAGE呈电泳均一的β亚基,Native-PAGE表明其为聚体,有弱的ATPase活性。The fragment of pGEM-T easyβ, containing the gene of chaperonin subunit β of Acidianus tengchongensis strain S5, was inserted into the plasmid pET23b with NdeI and BamHI, designated as pET23bβ and transformed into BL21 （DE3） or Rosetta- gami^MTB （DE3） pLysS. The expressed protein was soluble. Expression of the subunit β was 16. 2% of total cell proteins in Rosetta-gami^TM B （DE3） pLysS and displayed both monomer and oligomer. The recombinant subunit β was purified by means of sonication, heating at 70℃ for 10 min, ammonium sulfate precipitation, chromatography on Bio-Gel A-1.5m and DEAE-Sepharose CL-6B. The purified recombinant of subunit β displayed oligomer on native-PAGE and exhibited weak ATPase activity.

关 键 词：嗜酸热古菌 腾冲嗜酸两面菌 分子伴侣 Β亚基 ATPASE 

分 类 号：Q936[生物学—微生物学]