Glutamate déshydrogénase

Abstract

The complexes which mammalian‐liver glutamate dehydrogenase forms with its coenzymes, substrates and effectors have been characterised by difference spectroscopy, optical‐rotatory‐dispersion and circular‐dichroism studies. Particular attention was paid to the enzyme · NADH · glutamate complex which was studied in the absence and in the presence of the effectors ADP and GTP. Formation of the complexes is generally related both to modifications in the conformation of the enzyme and to the interaction between the enzyme and the coenzyme. Binding of the substrates to the NADH · enzyme complex induces changes in the circular‐dichroism spectrum of strates to the NADH · enzyme complex induces changes in the circular‐dichroism spectrum of the bound coenzyme: 2‐oxoglutarate increases the amplitude of the signal and glutamate an inversion of the spectrum. The amount of the non‐productive complex enzyme · NADH · glutamate formed during the catalysis is very high as long as the concentration of 2‐oxoglutarate remains low in comparison with the concentration of glutamate. There is one specific binding site for each ligand on each protomer ( M r = 56000) of the enzyme. Binding of the effectors induces no variation in the number of sites either of the coenzymes or of the substrates. Besides binding to its own active site, NADH may also bind to the adenylic site of ADP. However, formation of the ternary complex enzyme · NADH · glutamate induces an increase in the affinity of NADH for its active site and an important decrease in its affinity for the adenylic site. Competition for the catalytic site between NAD + and NADH and between L‐glutamate and 2‐oxoglutarate has been confirmed by rotatory power studies. It has been shown that GTP increases and ADP decreases the stability of the enzyme · coenzyme · substrate ternary complexes. GTP promotes the formation of the complex E · NADH · 2‐oxoglutarate while ADP promotes that of E · NADH · glutamate. These opposite effects of ADP and GTP are discussed in relation to the regulation of the enzymatic activity of glutamate dehydrogenase.