NAD-SPECIFIC GLUTAMATE-DEHYDROGENASE FROM THERMUS-THERMOPHILUS HB8 - PURIFICATION AND ENZYMATIC-PROPERTIES

An NAD-specific glutamate dehydrogenase from Thermus thermophilus HBS was purified 350-fold by a several-step procedure involving Blue-Sepha rose chromatography. The native protein had a molecular mass of approx imately 289 kDa, and consisted of six subunits with a molecular mass o f 48 kDa each. The optimum pH for-the deaminating reaction was 8.0. Th e optimum temperature was around 85-90 degrees C. NAD-glutamate dehydr ogenase showed a high coenzyme specificity, catalysed preferentially g lutamate catabolism and presented K-m values for NAD and L-glutamate o f 0.27 +/- 0.03 mM and 49 +/- 10 mM, respectively. No activity was det ected with NADH or NADPH, e-oxoglutarate and ammonia. Enzyme activity was found to be very stable at 72 degrees C. Differential scanning cal orimetry revealed a t(m) of 95 degrees C for denaturation. (C) 1998 Fe deration of European Microbiological Societies. Published by Elsevier Science B.V.