The metabolism of γ-hydroxyglutamate in rat liver I. Enzymic synthesis of γ-hydroxy-α-ketoglutarate from pyruvate and glyoxylate

Abstract

• 1.

  1. A new enzyme which catalyses the reversible condensation of pyruvate and glyoxylate to form γ-hydroxy-α-ketoglutarate was purified in excess of 120-fold from rat-liver acetone powder.

• 2.

  2. The enzyme was specific for glyoxylate and pyruvate. Condensation could not be demonstrated between other aldehydic acids and keto acids which were tested, distinguishing this enzyme from the formaldehyde and pyruvate condensing enzyme (2-oxo-4-hydroxybutyrate formaldehyde-lyase, EC 4.1.2.1) and the cleavage enzyme of γ-hydroxyglutamate (4-hydroxyglutamate glyoxylate-lyase). The apparent K_m values (at optimal pH, 7.5) for pyruvate, glyoxylate and γ-hydroxy-α-ketoglutarate were 10 mM, 0.43 nM and 1.33 mM, respectively. The equilibrium facors the biosynthesis of γ-hydroxy-α-ketoglutarate.

• 3.

  3. The product of the enzymic condensing reaction was isolated as the Ca salt and identified as γ-hydroxy-α-ketoglutaric acid from its chemical properties.

• 4.

  4. On the basis of the experimental findings, the possible metabolic pathways of pyruvate, glyoxylate and γ-hydroxyglutamate (the main oxidation product of L-hydroxyproline) and the regulating role of glyoxylate for the tricarboxylic acid cycle were discussed.