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• W1874448345 abstract "Glutamine-dependent carbamoyl-phosphate synthetase, the first enzyme of de novo pyrimidine nucleotide biosynthesis, was purified 1300-fold from Yoshida ascites hepatoma cells (AH 13) as a multi-enzyme complex with aspartate carbamoyltransferase and dihydroorotase, using dimethyl sulfoxide, glycerol and dithiothreitol as enzyme stabilizers. The ratios of the three enzyme activities were initially 1:65:9.2 and were essentially constant throughout the purification, and the final fraction had ratios of 1:37:11 (dihydroorotase assayed in the forward direction). The purified complex had a specific activity of the synthetase of 0.51 μmol × min−1× (mg protein)−1. It was essentially homogeneous on agarose/acrylamide composite gel electrophoresis, analytical ultra-centrifugation, and electrofocusing; it had an apparent pI of 5.5. The complex had a sedimentation coefficient (s20, w) of 24.6 S in the presence of 30% (v/v) dimethyl sulfoxide and 5% (w/v) glycerol, and its molecular weight was estimated to be 870000 by sedimentation equilibrium studies. On polyacrylamide gel electrophoresis in sodium dodecyl sulfate the complex migrated as a single protein band with a molecular weight of 210000, indicating that the complex is composed of four to five subunits of similar size. Sucrose gradient centrifugation in the presence of 1.5 M sodium chloride partially dissociated the three enzyme activities. Limited proteolysis of the complex with pancreatic elastase caused its dissociation and made separable the three activities. The synthetase was recovered as a 197000-molecular-weight fragment and the carbamoyltransferase as a 145000-molecular-weight fragment. Dihydroorotase was initially recovered as a 197000-molecular-weight fragment and was further converted to a 43000-molecular-weight fragment. It appears difficult to reconcile these results with the view that the three enzyme activities are located on the same polypeptide chain." @default.
• W1874448345 created "2016-06-24" @default.
• W1874448345 creator A5000776443 @default.
• W1874448345 creator A5091641496 @default.
• W1874448345 date "1978-05-01" @default.
• W1874448345 modified "2023-10-03" @default.
• W1874448345 title "Multi-Enzyme Complex of Glutamine-Dependent Carbamoyl-Phosphate Synthetase with Aspartate Carbamoyltransferase and Dihydroorotase from Rat Ascites-Hepatoma Cells. Purification, Molecular Properties and Limited Proteolysis" @default.
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• W1874448345 doi "https://doi.org/10.1111/j.1432-1033.1978.tb12320.x" @default.
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