重组人θ类谷胱甘肽转移酶（GSTT2-2）的纯化与鉴定
Purification and characterization of a recombinant human Theta-class glutathione transferase (GSTT2-2).
作者信息
Tan K L, Board P G
机构信息
Division of Molecular Medicine, John Curtin School of Medical Research, Australian National University, Canberra, Australia.
出版信息
Biochem J. 1996 May 1;315 ( Pt 3)(Pt 3):727-32. doi: 10.1042/bj3150727.
Abstract
A cDNA encoding the human Theta-class glutathione transferase GSTT2-2 was expressed in Escherichia coli as a ubiquitin fusion protein. The co-translational removal of the ubiquitin by a cloned ubiquitin-specific protease, Ubp1, generates enzymically active GSTT2-2 without any additional N-terminal residues. The recombinant isoenzyme was purified to apparent homogeneity by DEAE anion-exchange, gel filtration, dye ligand chromatography and high resolution anion-exchange chromatography on Mono Q FPLC. The recombinant enzyme had significant activity with a range of substrates, including cumene hydroperoxide and 1-menapthyl sulphate. The activity of GSTT2-2 with a range of secondary lipid peroxidation products such as the trans,trans-alka-2,4-dienals and trans-alk-2-enals, as well as its glutathione peroxidase activity with organic hydroperoxides, suggest that it may play a significant role in protection against the products of lipid peroxidation.
摘要
编码人θ类谷胱甘肽转移酶GSTT2-2的cDNA在大肠杆菌中作为泛素融合蛋白表达。通过克隆的泛素特异性蛋白酶Ubp1共翻译去除泛素，产生了具有酶活性的GSTT2-2，且没有任何额外的N端残基。通过DEAE阴离子交换、凝胶过滤、染料配体色谱和Mono Q FPLC上的高分辨率阴离子交换色谱将重组同工酶纯化至表观均一。重组酶对一系列底物具有显著活性，包括氢过氧化异丙苯和1-萘基硫酸盐。GSTT2-2对一系列次级脂质过氧化产物（如反式、反式-alka-2,4-二烯醛和反式-alk-2-烯醛）的活性，以及其对有机氢过氧化物的谷胱甘肽过氧化物酶活性，表明它可能在抵抗脂质过氧化产物方面发挥重要作用。
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