Purification, characterization and crystallization of pyrroline-5-carboxylate reductase from the hyperthermophilic archeon Sulfolobus Solfataricus
Purification, characterization and crystallization of pyrroline-5-carboxylate reductase from the hyperthermophilic archeon Sulfolobus Solfataricus
复制标题
超嗜热古细菌硫磺叶菌中吡咯啉-5-羧酸还原酶的纯化、表征和结晶
DOI：
10.1016/j.pep.2008.10.018
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发表时间：
2009-04-01
影响因子：
1.6
通讯作者：
Rao, Zihe
中科院分区：
文献类型：
--
作者：
Meng, Zhaohui;Liu, Zhe;Rao, Zihe
克隆并表达了嗜热嗜酸菌Sulfolobus solfataricus P2（Ss-P5 CR）的吡咯啉-5-羧酸还原酶（P5 CR）基因SSO 0495（proC）。纯化的重组酶催化硫代脯氨酸脱氢酶，同时将NAD（P）H氧化为NAD（P）（+）。这种古生菌酶在这种可逆反应中具有最佳碱性pH值，并且是热稳定的，在80 ℃下的半衰期约为30分钟。在pH 9.0时，反向活化速率比pH 7.0时高近3倍。均聚物的特征在于通过交联和尺寸排阻凝胶过滤色谱。通过悬滴气相扩散法在37 ℃下使Ss-P5 CR结晶。衍射数据获得的分辨率为3.5埃，适合于X射线结构测定。(C)2008年爱思唯尔公司All rights reserved.
The gene SSO0495 (proC), which encodes pyrroline-5-carboxylate reductase (P5CR) from the thermoacidophilic archeon Sulfolobus solfataricus P2 (Ss-P5CR), was cloned and expressed. The purified recombinant enzyme catalyzes the thioproline dehydrogenase with concomitant oxidation of NAD(P)H to NAD(P)(+). This archeal enzyme has an optimal alkaline pH in this reversible reaction and is thermostable with a half-life of approximately 30 min at 80 degrees C. At pH 9.0, the reverse activation rate is nearly 3-fold higher than at pH 7.0. The homopolymer was characterized by cross-linking and size exclusion gel filtration chromatography. Ss-P5CR was crystallized by the hanging-drop vapor-diffusion method at 37 degrees C. Diffraction data were obtained to a resolution of 3.5 angstrom and were suitable for X-ray structure determination. (C) 2008 Elsevier Inc. All rights reserved.