Male, K.B. and Storey, K.B. 1983. Tissue specific isozymes of glutamate dehydrogenase from the Japanese beetle, Popillia japonica: catabolic versus anabolic GDH's. J. Comp. Physiol. B 151, 199-205.

Tissue Specific Isozymes of Glutamate Dehydrogenase from the Japanese Beetle, Popillia japonica: Catabolic vs Anabolic GDH's

Keith B. Male and Kenneth B. Storey

1. Glutamate dehydrogenase (GDH) from the Japanese beetle, Popillia japonica, occurs in tissue specific isozymic forms. Two forms, specific for flight muscle and fat body, were identified and were separable by starch gel electrophoresis and by differential elution from NAD-agarose.

2. The isozymes utilized both NAD(H) and NADP(H) as coenzymes with activity ratios NADH: NADPH of 6: 1 for flight muscle and 8: 1 for fat body. pH optima for both enzymes were similar.

3. GDH from the two tissues differed kinetically. Affinity for alpha-ketoglutarate was much higher for the fat body enzyme, SO 5 for the NADH and NADPH linked reactions being 0.81+ 0.09 and 0.26+0.03 mM for fat body and 2.4+0.03 and 2.3 + 0.4 mM for flight muscle GDH, respectively.

4. Flight muscle GDH was much more strongly regulated by nucleotides than was the fat body isozyme. The apparent aestivation constant, Ka for ADP was 2-3 fold lower for the flight muscle enzyme for both forward and reverse reactions and ADP had a greater effect in lowering SO 5 for NH4+ for flight muscle GDH. GTP was a strong inhibitor of flight muscle GDH with apparent inhibitor constants, I50, of 15.5+3.0, 4.0+0.9 and 6.5+0.9 microM for the NADH, NADPH and NAD linked reactions, respectively. Fat body GDH, however, was only weakly affected by GTP with an I50 of 60+6 microM for the NAD reaction and I50's of greater than 500 microM for the NADH and NADPH linked reactions.

5. The kinetic properties of the two GDH isozymes suit the probable roles of the enzyme in vivo. Flight muscle GDH has a major role in the oxidation of proline as a fuel for flight. Nucleotide control of GDH would allow enzyme activity to respond to the energy status of the cell and would achieve a rapid activation of GDH at the initiation of flight. Fat body GDH, however, has a major role in the biosynthesis of proline and other amino acids. Enzyme activity is probably regulated by substrate availability, the absence of strong nucleotide regulation allowing enzyme function in high energy, resting states.