PURIFICATION AND PROPERTIES OF MALATE-DEHYDROGENASE FROM PARACOCCUS-DENITRIFICANS

Affinity chromatography on immobilized Cibacron Blue (Matrex Gel Blue A) gel permeation chromatography on UltroPac TSK G 3000 SWG column and ion-exchange chromatography on ''Mono Q'' column were used to purify the malate dehyhrogenase (MDH) from P. denitrificans to electrophoreti c homogeneity. The last two purification steps were performed in FPLC system. The enzyme having a specific activity of about 2300 nkat/mg pr otein was obtained with an approximate 70% yield. MDH is a dimer with a molecular mass of 80000 +/- 10000 and an isoeletric point of 4.85 +/ - 0.05. Absorption, fluorescence and CD-spectra were also measured and basic kinetic parameters were obtained for the homogeneous enzyme. Th e present paper also suggests the possibility of using the prepared en zyme for the determination of aspartate transferase (AST) in blood ser um.