Isolation and Characterization of Arginine-inducible Acetylornithine δ-Transaminase from Escherichia coli

Arginine-inducible acetylornithine δ-transaminase, which catalyzes the reversible conversion of N-acetylglutamic γ-semialdehyde and l-glutamate to α-N-acetyl-l-ornithine and α-ketoglutarate, has been isolated in homogeneous form, as judged by disc gel electrophoresis, immunoelectrophoresis, and ultracentrifugation. A double mutant of Escherichia coli strain W which lacks arginine-repressible acetylornithine δ-transaminase was used as the source of aminotransferase. In the assay used, optimal enzymatic activity occurs at pH 7.8, and requires the addition of pyridoxal 5-phosphate. The activity was measured in the reverse of the biosynthetic reaction and, under this condition, only α-N-acetyl-l-ornithine and α-ketoglutarate were found to serve as substrates. A number of divalent cations tested do not affect the activity of the enzyme, with the exception of Cu++ and Hg++, which are inhibitory. From amino acid analysis, the enzyme contains 5 half-cystine residues per molecule, and a partial specific volume of 0.734 has been calculated. The average molecular weight of acetylornithine δ-transaminase, based upon sedimentation equilibrium data, is 61,000, and an s020,w of 4.6 has been determined.