Glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic archaebacterium Pyrococcus woesei: characterization of the enzyme, cloning and sequencing of the gene, and expression in Escherichia coli
Glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic archaebacterium Pyrococcus woesei: characterization of the enzyme, cloning and sequencing of the gene, and expression in Escherichia coli
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来自超嗜热古细菌沃氏火球菌的甘油醛-3-磷酸脱氢酶：酶的表征、基因的克隆和测序以及在大肠杆菌中的表达
DOI：
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发表时间：
1990
影响因子：
3.2
通讯作者：
Reinhard Hensel
中科院分区：
文献类型：
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作者：
Peter Zwickl;S. Fabry;Christoph Bogedain;Andrea Haas;Reinhard Hensel
将来自超嗜热古细菌沃氏火球菌（Pyrococcus woesei）（最佳生长温度，100至103 ℃）的甘油醛-3-磷酸脱氢酶纯化至均一。这种酶是严格的磷酸盐依赖性，利用NAD+或NADP+，并不敏感pentalenolactone一样的产甲烷古细菌Methanothermus fervidus的酶。该酶表现出相当大的热稳定性，在100 ℃下半衰期为44分钟。根据沃氏毕赤酵母3-磷酸甘油醛脱氢酶编码基因的核苷酸序列推导出其氨基酸序列。与来自嗜温古细菌（Methanobacterium bryantii，Methanobacterium formicicicum）和极端嗜热古细菌（Methanothermus fervidus）的酶同源物相比，沃氏毕赤酵母酶的一级结构中芳香族氨基酸残基的比例非常高，而含硫残基的比例很低。沃氏毕赤酵母的编码基因在大肠杆菌中得到了高效表达，为进一步研究该酶的结构和功能奠定了基础。
The glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic archaebacterium Pyrococcus woesei (optimal growth temperature, 100 to 103 degrees C) was purified to homogeneity. This enzyme was strictly phosphate dependent, utilized either NAD+ or NADP+, and was insensitive to pentalenolactone like the enzyme from the methanogenic archaebacterium Methanothermus fervidus. The enzyme exhibited a considerable thermostability, with a 44-min half-life at 100 degrees C. The amino acid sequence of the glyceraldehyde-3-phosphate dehydrogenase from P. woesei was deduced from the nucleotide sequence of the coding gene. Compared with the enzyme homologs from mesophilic archaebacteria (Methanobacterium bryantii, Methanobacterium formicicum) and an extremely thermophilic archaebacterium (Methanothermus fervidus), the primary structure of the P. woesei enzyme exhibited a strikingly high proportion of aromatic amino acid residues and a low proportion of sulfur-containing residues. The coding gene of P. woesei was expressed at a high level in Escherichia coli, thus providing an ideal basis for detailed structural and functional studies of that enzyme.