Purification and characterization of a cold-adapted isocitrate lyase and a malate synthase from Colwellia maris, a psychrophilic bacterium.

Isocitrate lyase (ICL) and malate synthase (MS) of a psychrophilic marine bacterium, Colwellia maris, were purified to electrophoretically homogeneous state. The molecular mass of the ICL was found to be 240 kDa, composed of four identical subunits of 64.7 kDa. MS was a dimeric enzyme composed of 76.3 kDa subunits. N-Terminal amino acid sequences of the ICL and MS were analyzed. Purified ICL had its maximum activity at 20 degreesC and was rapidly inactivated at the temperatures above 30 degreesC, but the optimum temperature for the activity of MS was 45 degreesC. NaCl was found to protect ICL from heat inactivation above 30 degreesC, but the salt did not stabilize MS. Effects of temperatures on the kinetic parameters of both the enzymes were examined. The K-m for the substrate (isocitrate) of ICL was decreased with decreasing temperature. On the other hand, the K-m for the substrate (glyoxylate) of MS was increased with decreasing temperature. The calculated value of free energy of activation of ICL was on the same level as that of MS.