Liu Tiehan, Liu Shuang-Jiang, Xue Yanfen, Ma Yanhe, Zhou Peijin
Institute of Microbiology, Chinese Academy of Sciences, Beijing, ROC.
Extremophiles. 2002 Apr;6(2):97-102. doi: 10.1007/s007920100223.
The extremely halophilic archaeon ZP-6 was isolated from Ai-Ding salt lake in Xinjiang Uighur Autonomous Region of the People's Republic of China. Based on its physiological properties, 16S rDNA sequence, and DNA-DNA homology with known haloarchaea, the isolate was tentatively identified as a Halobacterium sp. An acetoacetyl-CoA thiolase was purified and characterized from this organism. The native enzyme has a molecular mass of 80 +/- 8 kDa and consists of two identical subunits of 43 +/- 2 kDa each. The N-terminus 14 amino acid residues were sequenced and showed identity with the respective part of a putative thiolase (AcaB1) of Halobacterium sp. NRC-1. The purified enzyme has an optimal pH of 7.9 for acetoacetyl-CoA thiolysis. The thiolytic activity was inhibited by the presence of Mg'- and was stimulated by KCl or NaCl. The thiolysis reaction of Halobacterium sp. ZP-6 thiolase can be inhibited by either substrate when present in excess. The distinct kinetic profile indicates that the thiolase from Halobacterium sp. ZP-6 may have a different catalytic mechanism from the so-called ping-pong mechanism employed by other thiolases. To our knowledge, this is the first report of the purification and characterization of a halophilic thiolase from an archaeal species.
极端嗜盐古菌ZP-6是从中华人民共和国新疆维吾尔自治区艾丁盐湖分离得到的。根据其生理特性、16S rDNA序列以及与已知嗜盐古菌的DNA-DNA同源性，该分离物初步被鉴定为嗜盐杆菌属。从该生物体中纯化并表征了一种乙酰乙酰辅酶A硫解酶。天然酶的分子量为80±8 kDa，由两个相同的亚基组成，每个亚基的分子量为43±2 kDa。对N端的14个氨基酸残基进行了测序，结果显示与嗜盐杆菌属NRC-1的一种假定硫解酶（AcaB1）的相应部分具有同一性。纯化后的酶对乙酰乙酰辅酶A硫解作用的最适pH为7.9。硫解活性受到Mg²⁺的抑制，而受到KCl或NaCl的刺激。当底物过量存在时，嗜盐杆菌属ZP-6硫解酶的硫解反应会受到任何一种底物的抑制。独特的动力学特征表明，嗜盐杆菌属ZP-6的硫解酶可能具有与其他硫解酶所采用的所谓乒乓机制不同的催化机制。据我们所知，这是首次报道从古菌物种中纯化和表征嗜盐硫解酶。
