Aoshima M, Yamagishi A, Oshima T
Department of Molecular Biology, Tokyo University of Pharmacy and Life Science, Japan.
Arch Biochem Biophys. 1996 Dec 1;336(1):77-85. doi: 10.1006/abbi.1996.0534.
A gene coding for isocitrate dehydrogenase (ICDH) was cloned from a hyperthermophilic archaebacterium, Caldococcus noboribetus, and sequenced. The gene was preceded by a promoter-like sequence and was followed by a terminator-like sequence. The deduced amino acid sequence of C. noboribetus ICDH showed high similarities to eubacterial ICDH. In particular, extremely high identity scores were found for ICDHs from Vibrio sp. (48.2%) and Escherichia coli (47.9%). The gene was expressed in E. coli by connecting it with the T7 promoter. The molecular weight of the gene product was estimated to be 48,000, which is consistent with that calculated from the deduced amino acid sequence. The gene product showed NADP-dependent ICDH activity at 80 degrees C. While the host-derived ICDH was completely inactivated by treatment at 70 degrees C for 10 min, the ICDH from C. noboribetus showed much higher thermostability.
从嗜热古细菌诺氏热球菌（Caldococcus noboribetus）中克隆并测序了一个编码异柠檬酸脱氢酶（ICDH）的基因。该基因前有一个类似启动子的序列，后有一个类似终止子的序列。诺氏热球菌ICDH推导的氨基酸序列与真细菌ICDH具有高度相似性。特别是，与弧菌属（Vibrio sp.）（48.2%）和大肠杆菌（Escherichia coli）（47.9%）的ICDH具有极高的同一性得分。通过将该基因与T7启动子连接，在大肠杆菌中进行表达。基因产物的分子量估计为48,000，这与从推导的氨基酸序列计算得出的结果一致。该基因产物在80℃时表现出NADP依赖的ICDH活性。虽然宿主来源的ICDH在70℃处理10分钟后完全失活，但诺氏热球菌的ICDH表现出更高的热稳定性。
