Photoreversal-dependent release of thymidine and thymidine monophosphate from pyrimidine dimer-containing DNA excision fragments isolated from ultraviolet-damaged human fibroblasts
To elucidate the enzymatic excision-repair process operative on cyclobutane-type pyrimidine photodimers in human dermal fibroblasts, we have examined excised dimer-containing material recovered in the trichloroacetic acid soluble fraction from far-ultraviolet-irradiated (254 nm, 40 J m-2) and incubated (24 h) cell cultures. The excised DNA photoproducts were found in oligonucleotide fragments with an estimated mean chain length of approximately 3.7 bases. Exposure of these isolated excision fragments, labeled with (/sup 3/H)thymidine (dT), to a secondary, dimer-photoreversing fluence of far-UV (5.5 kJ m-2) resulted in the release of free dT and thymidine monophosphate (TMP). Photorelease of these two radioactive species was measured by high-performance liquid chromatography, with TMP being detected as the increase in dT following bacterial alkaline phosphatase treatment. These data imply that the photoliberated dT and TMP moieties were attached to the excision fragments solely by the cyclobutane ring of the dimer. No evidence was obtained for the photoliberation of free thymine, thus corroborating a conclusion reached by others that the excision of dimers in human cells is not initiated by scission of an intradimer N-glycosyl bond. The sum of the tritium label recovered in dT plus TMP corresponded to approximately 40% of that disappearing from thymine-containing dimers on photoreversal, suggesting that in about 80% of the isolated excision fragments the dimer is located at one end of the oligonucleotide and contains a break in its internal phosphodiester bond.
- OSTI ID:
- 5120850
- Journal Information:
- Biochemistry; (United States), Vol. 9
- Country of Publication:
- United States
- Language:
- English
Similar Records
Dithymine photodimers and photodecomposition products of thymidylyl-thymidine induced by ultraviolet radiation from 150 to 300 nm
Processing of UV-induced DNA damage in diverse biological systems
Related Subjects
DNA REPAIR
BIOLOGICAL PATHWAYS
DNA
FIBROBLASTS
PHOTOREACTIVATION
PYRIMIDINE DIMERS
RADIOCHROMATOGRAPHY
SKIN
THYMIDINE
TRACER TECHNIQUES
TRITIUM COMPOUNDS
ULTRAVIOLET RADIATION
XERODERMA PIGMENTOSUM
ANIMAL CELLS
AZINES
BIOLOGICAL RECOVERY
BIOLOGICAL REPAIR
BODY
CHROMATOGRAPHY
CONNECTIVE TISSUE CELLS
DISEASES
ELECTROMAGNETIC RADIATION
HETEROCYCLIC COMPOUNDS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
NUCLEIC ACIDS
NUCLEOSIDES
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANS
PYRIMIDINES
RADIATIONS
RECOVERY
REPAIR
RIBOSIDES
SEPARATION PROCESSES
SKIN DISEASES
SOMATIC CELLS
560111* - Radiation Effects on Biochemicals- In Vitro- (-1987)