Partial purification and properties of carbamoyl phosphate synthetase of Alaska pea (Pisum sativum L. cultivar Alaska). Purification and general properties

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Abstract
1. Carbamoyl phosphate synthetase was purified up to 45-fold from Alaska pea seedling (Pisum sativum L. cultivar Alaska). 2. The enzyme was most active with and had the lowest Km for l-glutamine as compared with NH4+. 3. The purest preparations utilized very poorly or not at all l-asparagine and urea as nitrogen donors. 4. At saturating concentrations of components of the reaction, the Km for l-glutamine was 1·2×10−4m, and the Km for ATP was approx. 3·9×10−4m. 5. Although the enzyme was very labile, stability was improved by glutamine, asparagine, ammonium sulphate, dithiothreitol and especially l-ornithine. 6. Free ATP was markedly inhibitory, and MgATP2− and Mg2+ appeared to be the actual substrates utilized. 7. Fe2+ and Mn2+ were also utilized, but not as readily as Mg2+ except at low concentrations. K+ increased activity significantly. 8. Of the four nucleotides tested (ITP, ATP, GTP and UTP) only ATP served as an effective phosphate donor.