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《植物学报(英文版)》 2003-01
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Expression of Mouse MT-1 as a Fusion Protein in Anabaena sp. PCC 7120

ZHOU Jie, HAO Fu-Ying, SHI Ding-Ji, YU Mei-Min, RU Sing-Gen(1 . State Key- Laboratory of Protein Engineering and Plant Genetic Engineering, College of Life Sciences , Peking University, Beijing 100871, China; 2. Institute of Botany, The Chinese Academy of Sciences, Beijing 100093, China)  
To produce mouse metallothionein-1 (mMT-I ) in cyanobacterium Anabaena sp. PCC 7120, a novel Escherichia co/i-cyanobacterium shuttle fusion expression vector, pKG-MT, was constructed. Via this vector, mMT-I cDNA which was fused with a carboxyl terminal extension of the 26 kD glutathione-S-trans-ferase (GST) containing a thrombin specific site was expressed in Anabaena under the control of tac promoter. SDS-polyacrylamid gel electrophoresis (SDS-PAGE) showed that the fusion protein GST-MT was expressed in the transgenic Anabaena sp. PCC 7120 after induction with isopropylthio-β-D-galactoside (IPTG). Glutatione-S-transferase metallothionein (GST-MT) was purified from the crude extracts by affinity chromatography on immobilized glutathione and mMT- I was obtained by digesting the fusion protein with thrombin on column and gel filtration on Sephadex G-50. SDS-PAGE demonstrated that the purified mMT- I was the desired protein. The result of ELISA for the purified mMT-I showed that the recovery of mMT- I from the transgenic cyanobacterium was about 0.6 mg/g fresh weight. According to the data of atomic absorption assay, metal-binding activity of the purified mMT-I was almost the same as that of wild type MT.
【Fund】: 国家“九五”科技攻关计划(96-C02-04-05)~~
【CateGory Index】: Q786
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