Carbamoyl phosphate synthetase I of human liver. Purification, some properties and immunological cross-reactivity with the rat liver enzyme.
The purification of mitochondrial carbamoyl phosphate synthetase I (carbon-dioxide: ammonia ligase (ADP-forming, carbamate-phosphorylating), EC 6.3.4.16) from small samples of human liver is described. The enzyme is composed of a single polypeptide of Mr 160 000 +/- 500 as shown by SDS-polyacrylamide gel electrophoresis in the presence of reducing agents. The synthetase migrates in polyacrylamide gradient gels in the absence of detergents at a rate corresponding to a Mr of 165 000. Estimates of the molecular weight of the native enzyme by gel filtration and density gradient centrifugation yield a value of 178 000. The results indicate that the enzyme exists predominantly as monomeres. Amino acids composition, isoelectric point, stability, Km values and the ability to catalyze partial reactions have been measured and compared with known properties of carbamoyl phosphate synthetases from other sources. From the available data a high degree of evolutionary conservation of the ammonia-dependent synthetase is suggested. This is also supported by the demonstration of extensive immunological cross-reactivity between the human and rat enzymes.
D000596:Amino Acids / Q000032:analysis; D000645:Ammonium Sulfate; D000818:Animals; D001001:Anura; D002222:Carbamoyl-Phosphate Synthase (Ammonia) / Q000302:isolation & purification* / Q000378:metabolism; D000077286:Cetrimonium; D002593:Cetrimonium Compounds; D002852:Chromatography, Ion Exchange; D003429:Cross Reactions; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007231:Infant, Newborn; D008025:Ligases / Q000302:isolation & purification*; D008099:Liver / Q000201:enzymology*; D008970:Molecular Weight; D011817:Rabbits; D051381:Rats; D013045:Species Specificity
D000596:Amino Acids; D002593:Cetrimonium Compounds; D003902:Detergents; D008025:Ligases; D002222:Carbamoyl-Phosphate Synthase (Ammonia); D000645:Ammonium Sulfate; D000077286:Cetrimonium