Synthesis and structure-activity relationships of site-specific cis-syn and trans-syn thymine dimers
Upon UV irradiation of DNA, photoproducts are formed leading to mutations. The cis-syn and trans-syn thymine dimers are two such photoproducts. Lack of pure, well characterized lesions for study has hampered determination of biological structure-activity relationships. In this research, cis-syn and trans-syn thymine dimers have been placed site-specifically into DNA fragments for replication studies and into bacteriophage DNA for mutation studies. Although past studies on inhomogeneous templates indicate that cyclobutane dimers are absolute blocks to replication, we have determined that cis-syn cyclobutane dimers could be bypassed by several polymerases. A detailed study of E. coli polymerase I has indicated that trans-lesion synthesis of the cis-syn thymine dimer was dependent on time, dNTP and enzyme concentration. It is clear that the enzyme often dissociates upon replication into the 3{prime} thymine of the dimer. The binding constant of the enzyme to termination products is an important factor in bypass. Those enzymes that are most processive and bind tightest to the lesioned template, are able to bypass the dimer to the greatest extent. Addition of a processivity factor in one system enabled bypass of the trans-syn thymine dimer. High processivity is necessary, but not sufficient, for bypass of UV dimers. We have also examined the specificity of nucleotide insertion opposite the dimer site and have found the cis-syn dimer to be bypassed non-mutagenically. Although all nucleotides are inserted opposite the 3{prime} cis-syn dimer site to some extent, primers terminating in adenine are preferentially elongated by a series of selective kinetic steps. Several implications for the mechanisms by which polymerases are able to bypass the dimers are inferred from these studies.
- Research Organization:
- Washington Univ., Seattle, WA (United States)
- OSTI ID:
- 5543829
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
Cells
& Tissue Culture
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
AZINES
BACTERIA
BACTERIOPHAGES
BIOSYNTHESIS
CHEMICAL RADIATION EFFECTS
CHEMISTRY
DIMERS
DNA
DNA REPLICATION
ELECTROMAGNETIC RADIATION
ENZYMES
ESCHERICHIA COLI
HETEROCYCLIC COMPOUNDS
HYDROXY COMPOUNDS
MICROORGANISMS
MUTATIONS
NUCLEIC ACID REPLICATION
NUCLEIC ACIDS
NUCLEOTIDYLTRANSFERASES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PARASITES
PHOSPHORUS-GROUP TRANSFERASES
POLYMERASES
PYRIMIDINES
RADIATION CHEMISTRY
RADIATION EFFECTS
RADIATIONS
RADIOINDUCTION
STRUCTURE-ACTIVITY RELATIONSHIPS
SYNTHESIS
THYMINE
TIME DEPENDENCE
TRANSFERASES
ULTRAVIOLET RADIATION
URACILS
VIRUSES