Eubacteria-type isocitrate dehydrogenase from an archaeon: cloning, sequencing, and expression of a gene encoding isocitrate dehydrogenase from a hyperthermophilic archaebacterium, Caldococcus noboribetus

Abstract

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.