Glutamate dehydrogenase (L-glutamate: NAD+ oxidoreductase (deaminating), EC 1.4.1.2) has been purified 1250-fold from pea roots. The preparation contains only a single protein, and the molecular weight was estimated to be 208 000 ± 10 000. The enzyme shows NADH (aminating) and NAD+ (deaminating) activities, but the ratio of these activities is not constant and can be changed experimentally. NADPH activity is also present and shows a relatively constant ratio to NAD+ activity. EDTA inhibits NADH activity in intermediate concentrations, but reactivates at higher concentrations. NAD+ (and NADPH) activity is only slightly changed by EDTA. The effects of dioxane and the coenzymes on the enzyme are also reported. Mechanisms which could explain the different activity ratios, in terms of two interconvertible enzyme forms, are discussed.The pH optimum for NADH and NAD+ activities is about pH 8.0. Michaelis constants were found to be: α-ketoglutarate, 3.3 × 10−3 M; ammonium (sulfate), 3.8 × 10−2 M; glutamate, 7.3 × 10−3 M; NADH, 8.6 × 10−4 M; NAD+, 6.5 × 10−4 M. The enzyme is highly specific for the substrates glutamate and α-ketoglutarate, showing no alanine or aspartate dehydrogenase activity, and no deamination with a range of amino acids.
The regulation of activity of the enzymes involved in the assimilation of nitrate by higher plants
