Cloning, Sequencing, Expression and Characterization of the Gene Encoding the 3- Phosphoglycerate Kinase- Triosephosphate Isomerase Fusionprotein from Thermotoga Maritima

Abstract

Abstract: The gene encoding 3-phosphoglycerate kinase (PGK) and triosephosphate isomerase (TIM) from the hyperthermophilic bacterium Thermotoga maritima has been cloned in E.coli. The gene consists of two overlapping open reading frames. In E.coli, as in Thermotoga maritima, two proteins with corresponding N­ termini are expressed from this gene: a monomeric 43 kDa protein (PGK) and a tetrameric fusionprotein (PGKfus) of 280 kDa. The fusion protein reveals PGK and TIM activity and is synthesized as the result of a (-1)- frameshift. The nucleotide sequence was confirmed and the fusion protein was expressed and characterized. The recombinant enzymes are authenthic to those in Thermotoga maritima.