Glyoxysomal and mitochondrial malate dehydrogenase of watermelon (Citrullus vulgaris) cotyledons : II. Kinetic properties of the purified isoenzymes.

Kinetic parameters of the glyoxysomal and mitochondrial malate dehydrogenase (EC 1.1.1.37) of watermelon (Citrullus vulgaris Schrad.) cotyledons were compared. The data were obtained by initial rate experiments at pH 8.5 in both directions of the reaction using homogeneous enzyme preparations. Substrate inhibition at physiologically significant concentrations was observed with reduced nicotinamide adenine dinucleotide (NADH) (50% inhibition at 0.65 mmol·l(-1) NADH), but not with oxaloacetate, L-malate or oxidized nicotinamide adenine dinucleotide. The inhibition of both isoenzymes by 5'adenosine monophosphate was studied. Inhibition was found to be competitive with respect to NADH and non-competitive with respect to oxaloacetate. The apparent inhibitor constants at 200 μmol·l(-1) of the fixed substrates were 3.2 and 1.6 mmol·1(-1) for NADH, and 3.2 and 5.2 mmol·l(-1) for oxaloacetate with the glyoxysomal and mitochondrial isoenzymes, respectively. The energy of activation was determined for oxaloacetate reduction by glyoxysomal (E a =3.14×10(4)J×mol(-1)) and mitochondrial (E a =4.10×10(4) J x mol(-1)) MDH from Arrhenius plots, which exhibited constant slopes throughout the range of thermal stability.Despite considerable structural differences, the results indicate very similar kinetic behaviour of the glyoxysomal and mitochondrial isoenzymes. The physiological significance of the data are discussed in relation to the gluconeogenic processes occuring in cotyledons during germination.