Purification and stabilization of a monomeric isocitrate dehydrogenase from Corynebacterium glutamicum

Monomeric isocitrate dehydrogenase was expressed in Corynebacterium glutami cum cells harboring pEK-icdES1, a plasmid carrying the gene for the enzyme. Two- to three-fold higher expression levels of the recombinant enzyme were observed in such cells when grown in fermenters, compared to those grown i n shaker incubators. The enzyme was purified to homogeneity by ammonium sul fate fractionation, Sephadex G-150 gel filtration, FPLC Mono Q anion-exchan ge chromatography, and affinity gel chromatography. Approximately 4 mg of 9 8% pure recombinant enzyme was obtained per liter of bacterial culture. Our results also include optimum buffer conditions for purification and storag e of the enzyme. (C) 1999 Academic Press.