Characterization, Properties and Crystallization of Malate Dehydrogenase from Streptomyces coelicolor

Abstract

Malate dehydrogenase (MDH) isolated from Streptomyces coelicolor, A3(2) strain, electrophoretically homogenous, was crystallized in the absence or in the presence of NADH or NADPH coenzymes by the hanging-drop vapor-diffusion method. For simple isolation in a sufficient yield, the above MDH was cloned and expressed in Escherichia coli. Recombinant MDH with His-tag sequence did not crystallize whereas that without His-tag sequence crystallized using the above method. The crystals were grown in a mixture of 0.1 M sodium citrate buffer (pH 5.6), 0.2 M ammonium acetate, 27% (w/v) poly(ethylene glycol) 4000, and 5 mM NADH coenzyme or in a mixture of 0.1 M sodium citrate buffer (pH 4.8), 0.2 M ammonium acetate, 100 μM oxaloacetic acid, and 27% (w/v) poly(ethylene glycol) 4000.