CONTRASTING STRUCTURAL IMPACTS INDUCED BY CIS-SYN CYCLOBUTANE DIMER AND (6-4)-ADDUCT IN DNA DUPLEX DECAMERS - IMPLICATION LN MUTAGENESIS AND REPAIR ACTIVITY

Abstract

The relative biological importance of cis-syn cyclobutane dimer and pyrimidine(6-4)pyrimidone photoadduct ([6-4] photoadduct) appears to be dependent on the biological species, dipyrimidine sites and the local conformational variation induced at the damaged sites. The single-stranded deoxynucleotide 10-mers containing the site-specific (6-4) adduct or cis-syn cyclobutane dimer of thymidylyl (3'-->5')-thymidine were generated by direct photolysis of d(CGCAYTTACGC) with UVC (220-260 nm) irradiation or UVB (260-320 nm) photosensitization. Three-dimensional structures of the duplex cis-syn and (6-4) decamers of d(CGCATTACGC) d(GCGTAATGCG) were determined by NMR spectroscopy and the relaxation matrix refinement method. The NMR data and structural calculations establish that Watson-Crick base pairing is still intact at the cia-syn dimer site while the hydrogen bonding is absent at the 3'-side of the (6-4) lesion where the T --> C transition mutation is predominantly targeted. Overall conformation of the duplex cia-syn decamer was B-DNA and produced a 9 degrees bending in the DNA helix, but a distinctive base orientation of the (6-4) lesion provided a structural basis leading to 44 degrees helical bending. The observed local structure and conformational rigidity at the (6-4) adduct of the thymidylyl(3'-5')-thymidine (T-T [6-4]) lesion site suggest the potential absence of hydrogen bonding at the 3' sides of the (6-4) lesion with a substituted nucleotide during replication under SOS conditions. Contrasting structural distortions induced by the T-T (6-4) adduct with respect to the T-T cis-syn cyclobutane pyrimidine photodimer may explain the large differences in mutation spectrum and repair activities between them.