Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:6.3.5.5 (CPS)
1,262 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To study the regulation of transcription of the carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase (CAD) gene from the Syrian hamster, Mesocricetus auratus, we developed a homologous in vitro transcription system on the basis of nuclear extract from Syrian hamster kidney cells. We optimized the reaction temperature and the concentrations of DNA template, KCl, and MgCl2 simultaneously with the response surface method and found an unusually low temperature optimum of 20 degrees C. We therefore investigated whether CAD transcription in vitro depended on a heat-labile component of nuclear extract. Preincubating extract alone at 30 degrees C reduced transcription from the CAD promoter but not from the major late promoter of adenovirus 2. The formation of stable initiation complexes at the CAD promoter was diminished in heat-treated extract; run-off transcripts, however, accumulated at the same rate as in untreated extract. The heat sensitivity of complex formation correlated with the heat sensitivity of DNA binding by transcription factor Sp1, which binds to two sites in the CAD promoter; moreover, both preformed initiation complexes and DNA-bound Sp1 were heat-resistant. Adding purified Sp1 to heat-treated extract restored complex formation. We propose that Sp1 activates CAD transcription by stabilizing initiation complexes at the CAD promoter.
...
PMID:Heat sensitivity and Sp1 activation of complex formation at the Syrian hamster carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase promoter in vitro. 134 30

Carbamoyl-phosphate synthase II (glutamine-hydrolyzing) (EC 6.3.5.5) (synthase II) is the first and rate-limiting enzyme in the de novo UTP biosynthetic pathway. Leucine pulse-labeling in the rat demonstrated that in the rapidly proliferating hepatoma 3924A the ratio of radioactivity of synthase II to that of total cytosolic protein was 168.2 +/- 11.0 (SE) X 10(-3). This synthetic rate for the tumor enzyme was 9.7-fold higher than that for the liver synthase II, 17.4 +/- 4.0 X 10(-3). Since the degradation rate for hepatoma 3924A enzyme (t1/2 = 65.5 h) was similar to the rate for liver synthase II (t1/2 = 69.3 h), the increase in tumor synthase II activity and amount was due primarily to an elevation in enzyme synthesis in the presence of an unaltered catabolic rate. The results indicate that the reprogramming of gene expression in the hepatoma entails an increased production rate of the rate-limiting enzyme of UTP synthesis. This increase in the activity, concentration, and synthesis of tumor synthase II should provide a heightened capacity for the de novo pyrimidine biosynthetic pathway, thus conferring a selective advantage to the cancer cells.
...
PMID:Increased synthesis of carbamoyl-phosphate synthase II (EC 6.3.5.5) in hepatoma 3924A. 351 20

A high specific activity of carbamoyl-phosphate synthetase II (glutamine-hydrolyzing; EC 6.3.5.5) was demonstrated in extract of the cultured Crithidia fasciculata. The enzyme was separated from aspartate carbamoyltransferase by ammonium sulfate fractionation. Apparent Km for the synthetase for L-glutamine, NH4+, MgATP or bicarbonate was 0.27, 26, 1.7 or 1.7 mM at 2.0% dimethyl sulfoxide plus 0.3% glycerol. 8.6% dimethyl sulfoxide plus 1.4% glycerol decreased Km for L-glutamine to 0.10 mM, while Km for MgATP was unaffected. The higher solvent concentrations made Vmax markedly reduced, yielding the inhibition of the activity. These properties are unique to the Crithidia synthetase, compared with the mammalian enzyme.
...
PMID:Kinetic properties of carbamoyl-phosphate synthetase II (glutamine-hydrolyzing) in the parasitic protozoan Crithidia fasciculata and separation of the enzyme from aspartate carbamoyltransferase. 360 29

The effect of the anti-tumor, anti-glutamine drug acivicin, L-(alpha S,5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid, was determined on the activity of the rate-limiting enzyme of de novo pyrimidine biosynthesis, carbamoyl-phosphate synthetase II (glutamine-hydrolyzing) (EC 6.3.5.5), in human colon carcinoma. The synthetase II activity in human colon carcinoma was elevated 2- to 3-fold over values of the normal colon mucosa, and the substrate kinetic constants were similar for the enzyme in normal and neoplastic colon. The Km for glutamine was 17 microM (colon carcinoma) and 23 microM (normal mucosa), whereas the Km for ATP was 2.1 and 1.7 mM in tumor and mucosa respectively. The synthetase II activity in colon carcinoma was inhibited to a similar extent by UMP, UDP and UTP (36-41%). The three uracil nucleotides were also equally effective in inhibiting the enzyme from normal mucosa (39-46%). Both enzymes were activated by PRPP (63 and 57%) in mucosa and carcinoma respectively. Acivicin in vitro selectively inactivated the glutamine-dependent synthetase II from human colon carcinoma, and it did not affect the ammonia-dependent activity. The acivicin inactivation constant (Kinact) was 100 microM, and the minimum inactivation half-time (T) was 0.7 min. Acivicin most likely exerts its effect against human colon synthetase II by acting as an active site directed affinity analogue of L-glutamine.
...
PMID:Inactivation by acivicin of carbamoyl-phosphate synthetase II of human colon carcinoma. 396 20

Several recombinant plasmids containing cpaII, the gene that encodes the large subunit of yeast arginine-specific carbamoyl-phosphate synthetase [carbamoyl-phosphate synthetase (glutamine-hydrolyzing), carbon-dioxide: L-glutamine amido-ligase (ADP-forming, carbamate-phosphorylating), EC 6.3.3.5], have been isolated. The plasmids were selected by transformation of a yeast strain with a mutation in the structural gene of the large subunit of carbamoyl-phosphate synthetase. By using a recombinant pool with inserts of yeast nuclear DNA of 5-20 kilobase pairs, we obtained 13 transformants. Of five transformants studied, three have been found to have stable plasmid inserts. These plasmids could be amplified in Escherichia coli and transferred back into the yeast carbamoyl-phosphate synthetase-deficient strains with concomitant complementation of the nuclear mutation. Plasmids pJL2/T1 and pJL2/T5 contain identical nuclear DNA inserts of 5.9 kilobase pairs. Although the insert of plasmid pJL2/T3 is also 5.9 kilobase pairs long, the sequence overlap with pJL2/T1 and pJL2/T5 is only 4.5 kilobase pairs long. The T3 insert has an orientation in the vector opposite to that of the T1 and T5 inserts. The recombinant plasmids with the yeast cpaII gene fail to cross-hybridize with a cloned fragment of E. coli DNA containing the carA and carB genes for the bacterial carbamoyl-phosphate synthetase.
...
PMID:Cloning of a yeast gene coding for arginine-specific carbamoyl-phosphate synthetase. 628 75

Carbamoyl-phosphate synthetase II (glutamine-hydrolyzing) (EC 6.3.5.5) (synthetase II), is the first and rate-limiting enzyme in the de novo UMP biosynthetic pathway. The present investigation showed that insulin has a regulatory action on hepatic synthetase II activity. When diabetes was induced with injection of different doses of alloxan the plasma insulin concentrations decreased in a dose-dependent fashion to 72, 38, 31 and 28% and concurrently the liver synthetase II activity decreased to 75, 43, 29 and 22% of the normal values. In diabetic rats dose response studies showed that with insulin injections of 4, 6, 8 or 10 U/day for 48 h the hepatic synthetase II activity increased to 81, 95, 99 and 103% of the control liver values. In the diabetic rats the insulin-induced rise in liver synthetase II activity was prevented by treatment of the rats with actinomycin.
...
PMID:Action of insulin on liver carbamoyl-phosphate synthetase II (glutamine-hydrolyzing) activity. 634 26

In rat livers and hepatomas, carbamoyl phosphate synthetase (glutamine-hydrolyzing) (EC 6.3.5.5) (synthetase II), the rate-limiting enzyme of de novo pyrimidine nucleotide biosynthesis, was separated from carbamoyl phosphate synthetase (ammonia) (EC 6.3.4.16) (synthetase I) ammonium sulfate and hydroxylapatite fractionations and gel filtration on Sephadex G-25. Both liver and hepatoma 3924A synthetase II activities were subject to feedback inhibition by UTP and to stimulation by 5-phosphoribosyl 1-pyrophosphate. UTP (0.5 mM) enhanced the apparent Km for MgATP from 2.3 to 7.6 mM, whereas 0.1 mM 5-phosphoribosyl 1-pyrophosphate reduced it to 0.5 mM. At 2 mM MgATP, 3 or 7 microM 5-phosphoribosyl 1-pyrophosphate yielded half-maximal activation (Ka) in the absence or presence of 0.5 mM UTP; UTP altered the stimulation kinetics from hyperbolic to sigmoidal. In the rat, synthetase II activities were highest in thymus, testis and spleen. In differentiating and regenerating rat livers, activities were 2.2- and 1.5-fold higher than in adult livers. In 17 hepatomas of different growth rates, synthetase II activity increased 1.3- to 9.5-fold over liver values; the rise correlated positively with tumor growth rates. Synthetase II activities also increased in a kidney tumor (5.0-fold) and in a sarcoma (18.1-fold) in the rat and in a human colon tumor (3.3-fold).
...
PMID:Regulatory properties and behavior of activity of carbamoyl phosphate synthetase II (glutamine-hydrolyzing) in normal and proliferating tissues. 705 79

The specific activity of carbamoyl phosphate synthetase (glutamine-hydrolyzing), the first and rate-limiting enzyme of de novo uridine 5'-triphosphate biosynthesis, was increased in 13 transplantable hepatomas, particularly in the rapidly growing tumors (5.7- to 9.5-fold), and the rise was correlated with tumor growth rates. Thus, synthetase activity was linked with both hepatic neoplastic transformation and progression. Synthetase specific activity was so elevated in a transplantable sarcoma (18-fold) and a kidney adenocarcinoma (5-fold). The increased activity should enhance the capacity of the pathway and should confer selective advantages to cancer cells.
...
PMID:Carbamoyl phosphate synthetase (glutamine-hydrolyzing): increased activity in cancer cells. 720 43

To better understand the signaling pathways which lead to DNA synthesis in mammalian cells, we have studied the transcriptional activation of genes needed during the S phase of the cell cycle. Transcription of the gene encoding a pyrimidine biosynthetic enzyme, carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase (cad), increases at the G1/S-phase boundary. We have mapped the growth-dependent response element in the hamster cad gene to the extended palindromic E-box sequence, CCACGTGG, which is centered at +65 in the 5' untranslated sequence. Mutation of the E box abolished growth-dependent transcription, and an oligonucleotide corresponding to the cad sequence at +55 to +75 (+55/+75) restored growth-dependent regulation to nonresponsive cad promoter mutants when placed down-stream of the transcription start site. The same oligonucleotide conferred less G1/S-phase induction when placed upstream of basal promoter elements. An analogous oligonucleotide containing the mutant E box had no effect in either location. Nuclear proteins bound the cad +55/+75 element in a cell cycle-dependent manner in electromobility shift assays; antibodies specific to USF and Max blocked the DNA-binding activity of different growth-regulated protein-DNA complexes. Expression of c-Myc mutants which have been shown to dominantly interfere with the function of c-Myc and Max significantly inhibited cad transcription during S phase but had no effect on transcription from another G1/S-phase-activated promoter, dhfr. These data support a model whereby E-box-binding proteins activate serum-induced transcription from the cad promoter at the G1/S-phase boundary and suggest that a Max-associated protein complex contributes to the serum response.
...
PMID:An E-box-mediated increase in cad transcription at the G1/S-phase boundary is suppressed by inhibitory c-Myc mutants. 773 36

Transcription of the carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase (CAD) gene from the Syrian hamster, Mesocricetus auratus, starts at a single major site. We characterized the cis-acting elements that position RNA polymerase II at the correct start site in the CAD promoter. Sequence alignment showed that the CAD promoter lacks a TATA box, but contains a consensus initiator. Mutational analysis of the CAD promoter demonstrated that the sequences between -81 and +26 were sufficient for accurate and efficient transcription in vitro and in vivo; binding sites for the transcription factor Sp1 around -70 and -49 were necessary for transcriptional activity. The binding site at -49 directed initiation about 50 base pairs downstream. A ubiquitous activator protein, Honk, bound to the CAD promoter between -30 and -12, but did not participate in start site selection. The sequences around +1, which contain the consensus initiator, contributed to promoter activity; however, the presence of a consensus initiator in this region was neither necessary nor sufficient for transcription. We concluded from these results that the Sp1 binding site at -49 substituted for the missing TATA box and played a major role in start site selection at the CAD promoter.
...
PMID:Start site selection at the TATA-less carbamoyl-phosphate synthase (glutamine-hydrolyzing)/aspartate carbamoyltransferase/dihydroorotase promoter. 790


1

---