Control of pyrimidine biosynthesis in the Ascaris ovary: regulatory properties of glutamine-dependent carbamoyl-phosphate synthetase and copurification of the enzyme with aspartate carbamoyltransferase and dihydroorotase.
Glutamine-dependent carbamoyl-phosphate synthetase, the first enzyme of the de novo biosynthetic pathway for pyrimidine nucleotides, was purified about twenty-fold from 105 000 x g supernatant of the Ascaris ovary homogenate. The enzyme activity was feedback-inhibited by UDP and UTP while it was stimulated by 5-phosphoribosyl 1-pyrophosphate. Most of the catalytic and regulatory properties of the Ascaris synthetase were similar to those of the mammalian synthetase. A significant difference is that the Ascaris enzyme was more strongly inhibited by UDP than by UTP whereas the mammalian enzyme is more sensitive to UTP than to UDP. The Ascaris enzyme was also inhibited by other various nucleoside diphosphates, such as dUDP, dADP and CDP, generally more strongly than by the corresponding nucleoside triphosphates. Aspartate carbamoyltransferase and dihydroorotase, the second and third enzymes of the pathway, were also demonstrated in the supernatant fraction. These two enzymes were copurified with the synthetase and the relative activities of the three enzymes remained nearly constant (1:850-890:50-60) throughout the purification. In a sucrose gradient centrifugation, the enzymes cosedimented as a single peak with a sedimentation coefficient (s20,w) of about 32 S under the condition used. These results strongly suggest that the enzymes exist as a multienzyme complex similar to those found in higher animals. The activity of the carbamoyltransferase was insensitive to nucleotides and related compounds. These results indicate that the synthetase plays a key role in the control of pyrimidine biosynthesis in the Ascaris ovary.
D000581:Amidohydrolases / Q000378:metabolism*; D000818:Animals; D001200:Ascaris / Q000201:enzymology*; D001221:Aspartate Carbamoyltransferase / Q000302:isolation & purification / Q000378:metabolism*; D002223:Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing) / Q000302:isolation & purification / Q000378:metabolism*; D004080:Dihydroorotase / Q000302:isolation & purification / Q000378:metabolism*; D005260:Female; D008025:Ligases / Q000378:metabolism*; D010053:Ovary / Q000201:enzymology; D011743:Pyrimidines / Q000096:biosynthesis*
D011743:Pyrimidines; D001221:Aspartate Carbamoyltransferase; D000581:Amidohydrolases; D004080:Dihydroorotase; D008025:Ligases; D002223:Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing)