Release of hepatic mitochondrial ornithine transcarbamylase into the circulation in D-galactosamine-treated rats. Identification of serum ornithine transcarbamylase as the intact form of the mitochondrial enzyme.
A single injection of D-galactosamine into rats caused acute liver cell injury, and the activity of ornithine transcarbamylase in the serum increased about 600-fold as compared with that in the normal serum. Some properties of the serum enzyme from galactosamine-treated rats have been studied together with those of the mitochondrial enzyme in liver. Both the enzyme activities gave similar pH profiles, showing an optimum of pH 8.5. Apparent Km values of the serum enzyme for ornithine under the standard conditions at pH 7.4 and pH 7.7 were 1.59 mM and 0.94 mM, respectively, and those of the mitochondrial enzyme were 1.69 mM and 0.97 mM, respectively. The Km value of the serum enzyme for carbamyl phosphate was 0.34 mM, which is similar to that of the mitochondrial enzyme. The mitochondrial enzyme was purified 78-fold to homogeneity with a 45% yield by ammonium sulfate fractionation, heat treatment, 2nd ammonium sulfate fractionation, and phenyl-Sepharose CL-4B and CM-Sephadex C-50 column chromatographies. The specific activity of the purified enzyme was 282 mumol of citrulline formed per mg of protein per min at 37 degrees C. The mitochondrial and serum enzymes have a molecular weight of 115,000 as determined by Sephacryl S-300 gel filtration. Antibody specific for the mitochondrial enzyme was raised, and the immunological properties of the serum enzyme were examined. In immunoinhibition experiments, a decrease of the serum enzyme activity as well as the mitochondrial enzyme was observed on increasing the amount of the antibody.
D000818:Animals; D005688:Galactosamine / Q000633:toxicity*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008297:Male; D008930:Mitochondria, Liver / Q000201:enzymology*; D008970:Molecular Weight; D009954:Ornithine Carbamoyltransferase / Q000097:blood* / Q000302:isolation & purification / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D013997:Time Factors
D005688:Galactosamine; D009954:Ornithine Carbamoyltransferase