Purification and some properties of two NADP+-specific isocitrate dehydrogenases from an obligately psychrophilic marine bacterium, Vibrio sp., strain ABE-1.

Abstract

Two isozymes of NADP+-specific isocitrate dehydrogenase [ICDH; EC 1.1.1.42] were confirmed to be present in an obligately psychrophilic marine bacterium, Vibrio sp., strain ABE-1, on the basis of the temperature-activity curve and electrophoretic mobilities. These isozymes were separated and purified about 170-fold for isozyme I (specific activity at 40 degrees C, 24.3 units/mg protein) and about 180-fold for isozyme II (specific activity at 20 degrees C, 59.2 units/mg protein), though the isozymes were still not homogeneous. The molecular weights of these isozymes determined by gel filtration were both about 85,000, but the properties of the isozymes were considerably different from each other. The thermostability of isozyme I resembled those of mesophiles, but isozyme II was extremely labile above 20 degrees C. NaCl affected the ICDH isozymes in different ways; the salt protected isozyme I from heat inactivation, but not isozyme II. Nevertheless it enormously enhanced the activity of isozyme II at low concentrations. Moreover, these ICDH isozymes showed different pH optima, Km values for isocitrate, susceptibilities to concerted inhibition by glyoxylate plus oxalacetate, and effects of 2-mercaptoethanol on their stabilities.