Activity of Liver Glutamine Transaminase toward L-Y -Glutamyl Hydrazones of aXeto Acids*

Rat liver glutamine transaminase acts on y-glutamyl hydrazones of a-keto acids to yield the corresponding L-amino acids and 3-hydroxy-tetrahydro-6-pyridazinone-3-carboxyllc acid; the latter compound undergoes nonenzymatic dehydration in acid to yield 1,4,5,6-tetrahydro-6-pyridazinone -3carboxylic acid. L-Amino acid oxidase (snake venom) also catalyzes the formation of 3-hydroxy-tetrahydro-6-pyridazinone-3-carboxylic acid (and ammonia) from y-glutamyl hydrazide. In the transaminase-catalyzed reaction in which glycine is formed from the y-glutamyl hydrazone of glyoxylate, no evidence was obtained for the formation of free glyoxylate during the reaction, nor did added free glyoxylate equilibrate with y-glutamyl hydrazone-bound glyoxylate. The relative rates of reaction observed with a series of y-glutamyl cr-keto acid hydrazones were similar to those of the corresponding glutamine-a-keto acid transamination reactions; in general, the rates with the y-glutamyl Lu-keto acid hydrazones were greater than those of the corresponding glutamine-a-keto acid transamination reactions. Possible reaction pathways for the enzymatic transformation of the y-glutamyl a-keto acid hydrazones have been deduced. Albizziin (L-a!-amino-fl-ureidoproprionic) acid was found to effectively replace glutamine in the glutamine-cr-keto acid transamination reaction; the high ultraviolet absorbance of the dehydrated product of the Lu-keto acid analog of albizziin was used as the basis for a highly sensitive assay for liver glutamine transaminase activity. Studies of the albizziinglyoxylate transamination reaction indicated that the enzyme exhibits ping-pong kinetics. The data suggest that the enzyme has binding sites for glutamine and a-keto acids which overlap. The action of the enzyme on y-glutamyla-keto acid hydrazones appears to involve initial binding of the glutamyl moiety at the glutamine site of the enzyme followed by transformations which are associated with movement of the attached a-keto acid moiety into the active site. The preparation and properties of the L-y-glutamyl hydrazones of a number of ar-keto acids are described.