Deoxyribonucleotide pool analysis: functional association of thymidylate synthase with the other enzymes of DNA biosynthesis in mammalian cells
Allosteric interaction between thymidylate synthase (TS) and the other enzymes of DNA biosynthesis was suggested from the authors observation that inhibitors of ribonucleotide reductase, topoisomerase of DNA polymerase-..cap alpha.. inhibit TS in intact S phase CHEF/18 cells, but not in their soluble extracts. In addition the authors observed that 4'-(9-acridinylamino)-methanesulfon-m-anisidide (m-AMSA), a poison of topoisomerase II, had similar effects on TS activity in mammalian cells. They have examined if the inhibitory effects of these antimetabolites on TS is due to the accumulation of thymidine nucleotide(s) in intact cells, rather than to an allosteric interaction in the replitase complex. A novel method of nucleotide pool analysis revealed that in the presence of these antimetabolites the incorporation of radioactivity from /sup 3/H-deoxyuridine (dUrd) into thymidine nucleotide pools inside the cell did not increase as compared to the control. Furthermore, TS activity as measured in-vitro was not inhibited by supraphysiological concentrations (50..mu..M) of thymidine mono- or tri-phosphates. None of these antimetabolites dramatically influenced the uptake of dUrd and its subsequent phosphorylation to deoxyuridine monophosphate. Therefore, they suggest that the inhibitory effect of these antimetabolites is due to the functional association of their target enzymes with TS.
- Research Organization:
- Univ. of Virginia, Charlottesville
- OSTI ID:
- 7189752
- Report Number(s):
- CONF-8606151-
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Vol. 45:6; ISSN FEPRA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
ANIMAL CELLS
ANIMALS
ANTIMETABOLITES
AZINES
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL EFFECTS
BIOSYNTHESIS
DEOXYURIDINE
DNA
DRUGS
ENZYME ACTIVITY
ENZYMES
HETEROCYCLIC COMPOUNDS
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
KINETICS
LABELLED COMPOUNDS
LIGASES
MAMMALS
METABOLISM
NUCLEIC ACIDS
NUCLEOSIDES
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
OXYGEN COMPOUNDS
PHOSPHATES
PHOSPHORUS COMPOUNDS
PYRIMIDINES
REACTION KINETICS
RIBOSIDES
SYNTHESIS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
URACILS
VERTEBRATES