PMID: 9555069Jun 20, 1998Paper

Studies on the regulation of CTP:phosphocholine cytidylyltransferase using permeabilized HEP G2 cells: evidence that both active and inactive enzyme are membrane-bound

Biochimica Et Biophysica Acta
P A Weinhold, D Barrett

Abstract

To obtain more insight into the mechanisms regulating CTP:phosphocholine cytidylyltransferase (CT), we determined the effect of oleate treatment on the rate of CT release from permeabilized Hep G2 cells and the distribution of the CT remaining in the permeabilized cells. When we permeabilized untreated cells in pH 7.5 buffer containing 0.15 M KCl, the rate of CT release was much slower than the release of lactate dehydrogenase. Oleate treatment caused a further decrease in CT release from cells. In untreated cells, 70-80% of the CT remaining in cells 10 min after permeabilization was recovered as soluble CT. Oleate treatment increased the amount of bound CT but over 50% of the CT in cells 10 min after permeabilization was recovered as soluble CT. In both control and oleate-treated cells, the increase in CT release with time correlated with a decrease in the amount of CT recovered from permeabilized cells as soluble CT. These results suggested that CT existed in a form that was not immediately available for release from permeabilized cells, but was recovered in the soluble fraction after cell disruption. When cells were permeabilized in 10 mM imidazole-20% glycerol-5 mM Mg2+ pH 6.5, over 80% of CT in control and over 90% of CT i...Continue Reading

References

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