Glutamate dehydrogenase (GDH) represents a critical enzyme catalyzing the reaction spanning amino acid catabolism, the Krebs cycle, and urea production in the wood frog. GDH breaks down glutamate and NAD+ to generate α-ketoglutaric acid (α-KG), NADH and ammonium that can be metabolized to form urea. Purification of GDH from control and frozen male wood frog livers was performed using a two-step column chromatography procedure with a cation exchange column and a GTP-agarose affinity column. Analysis of kinetic parameters of the purified GDH showed several notable differences between the control and stress. Under standard assay conditions, the affinity of GDH for its substrates was significantly higher for the freeze-exposed enzyme than for the control (glutamate Km: 41% decrease, NAD+ Km: 40% decrease). The maximal activity for the control enzyme was also noted to be lower than the frozen. This suggests that the frozen form of the GDH was activated relative to the control form. Western blot analysis of common posttranslational modifications indicated that the frozen enzyme had a lower degree of acetylation and ADP-ribosylation than its control counterpart. These results suggest that GDH is regulated in the wood frog liver by means of altering post-translational modifications in response to freezing.