Radiometric determination of cytidine 5′-triphosphate in biological materials

Leelavathi, D.E.; Guynn, Robert W.

doi:10.1016/0003-2697(77)90534-6

Cited by 5 publications

(4 citation statements)

References 16 publications

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“…The disadvantage of that procedure is that extensive purification of the enzyme is required. Our result for rat liver CTP is in agreement with the reported value for fasted rats of 0.047 prnollg liver (20).…”

Section: P P Bnyolisupporting

confidence: 93%

“…ported that CTP could be measured by an assay that used phosphoethanolamine cytidylyltmnsferase (20). The disadvantage of that procedure is that extensive purification of the enzyme is required.…”

Section: P P Bnyolimentioning

confidence: 99%

“…The advantages of the present method over published procedures for measuring CTP and phosphocholine (5,6,20) are the extreme ease of preparation of phosphocholine cytidylyltransferase, the separation of CTP and phosphocholine which enables one to make an independent measurement of the concentration of each compound from the same sample, and the sensitivity and specificity of an enzymatic determination. With the availability of this method, we are now in a position to test our hypothesis that the concentration of CTP may determine the rate of phosphatidylcholine biosynthesis in animal cells under certain physiological conditions.…”

Section: P P Bnyolimentioning

confidence: 99%

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A rapid method for the determination of CTP and phosphocholine in rat liver and baby hamster kidney 21 cells

Choy¹,

Whitehead²,

Vance³

1978

Can. J. Biochem.

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A rapid and sensitive assay for CTP and phosphocholine was required for us to determine the concentration of these compounds in tissues and cell cultures. Such a procedure was devised with CTP:phosphocholine cytidylyltransferase, an enzyme which is highly specific for CTP and phosphocholine. The 0--22% ammonium sulfate precipitate of a cytosolic extract from rat liver was used as the source of the enzyme. The amount of CTP in an extract was estimated by the conversion of [3H]phosphocholine to 3H-labelled CDP-choline. Similarly, the concentration of phosphocholine was estimated by the formation of 3H-labelled CDP-choline from 3H-labelled CTP. The conversion of CTP and phosphocholine to CDP-choline was 90% when inorganic pyrophosphatase was added to the incubations. The formation of CDP-choline was linear between 1 and 10 nmol of CTP or phosphocholine. The concentration of CTP was determined in rat liver (62 nmol/g wet weight) and baby hamster kidney 21 (BHK-21) cells (161 nmol/g wet weight). The concentration of phosphocholine in rat liver was 1.16 mumol/g wet weight whereas in BHK-21 cells it was much less (69 nmol/g wet weight). By this procedure, it may be possible to establish the importance of CTP and phosphocholine in the control of phosphatidylcholine biosynthesis.

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Section: P P Bnyolisupporting

confidence: 93%

Section: P P Bnyolimentioning

confidence: 99%

Section: P P Bnyolimentioning

confidence: 99%

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A rapid method for the determination of CTP and phosphocholine in rat liver and baby hamster kidney 21 cells

Choy¹,

Whitehead²,

Vance³

1978

Can. J. Biochem.

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“…Our analysis revealed that after 48 h of fasting, the reduction in phosphatidylcholine biosynthesis was caused by changes in several factors, including a diminishing rate of choline uptake, reductions in the pool sizes of ATP, CTP and cholinecontaining metabolites and alterations in activities of the enzymes in the CDP-choline pathway. The reduction in the high-energy nucleotide level in the liver during fasting has been documented (Leelavathi and Guynn, 1977). We have shown in the hypoxic heart (Hatch and Choy, 1990) and during the development of cardiomyopathies (Choy, 1982) that the impairment of phosphatidylcholine biosynthesis was a direct result of severe reductions in high-energy nucleotides levels.…”

Section: Discussionmentioning

confidence: 86%

Effects of fasting on phosphatidylcholine biosynthesis in hamster liver: regulation of cholinephosphotransferase activity by endogenous argininosuccinate

Choy

1993

Biochemical Journal

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The control of phosphatidylcholine biosynthesis in the hamster liver was examined. Livers of hamsters fasted for 24 and 48 h were perfused with labelled choline. Under both fasting conditions, the incorporation of labelled choline into phosphatidylcholine was reduced. After 48 h of fasting, the 52% reduction in phosphatidylcholine biosynthesis was caused by changes in several factors including a diminishing rate of choline uptake and severe reductions in the pool sizes of ATP and CTP (to 33-37% control values) which resulted in a decrease in the pools of choline-containing metabolites. The activation of cytidylyltransferase after 48 h of fasting might be regarded as a compensatory mechanism for the maintenance of phosphatidylcholine biosynthesis. After 24 h of fasting, a 25% reduction in phosphatidylcholine biosynthesis was observed. The ATP and CTP levels were decreased but the reduction was not severe enough to affect the choline uptake or the labelling of the phosphocholine fraction. The activities of the cytidylyltransferase remained unchanged but an accumulation of labelled CDP-choline was detected. Although choline-phosphotransferase activity was not changed in the microsomes, the enzyme activity was attenuated in the postmitochondrial fraction. Further analysis revealed that cholinephosphotransferase in the liver was inhibited by an endogenous inhibitor in the cytosol which was later identified as argininosuccinate. The level of argininosuccinate was elevated during fasting and the change quantitatively accounted for the attenuation of cholinephosphotransferase activity. The inhibition of choline-phosphotransferase by argininosuccinate, coupled with a substantial decrease in the diacylglycerol level, would provide the hamster liver with an immediate mechanism for the transient modulation of phosphatidylcholine biosynthesis during short-term fasting.

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