Abasic site residues

The abasic sites (3, Scheme 8.2) resulting from the loss of alkylated bases from DNA are both cytotoxic and mutagenic. " The cyclic acetal (3) exists in equilibrium with small amounts (—1%) of the open chain aldehyde (4). The acidic nature of a-proton in the aldehyde form of the abasic lesion facilitates 3-elimination of the 3 -phosphate residue to yield a strand break. " This reaction occurs with a half-life of about 200 h under physiological conditions (pH 7.4, 37°C), but can be accelerated by heat, basic conditions, or the presence of various amines. " ... 

Besides the direct oxidation products of base residues of DNA, such as the oxidation of thymine (43) and guanine (160) residues meutioued in Sections IV.B.2 and IV.B.3, ROS and other factors may bring about detachmeut of base residues from the DNA strand, leading to formation of aldehyde abasic sites on the main chain, as shown in... 

E. coli Nth protein (endonuclease III) Oxidized pyrimidines Ring-saturated or fragmented Thy residues, e.g. F Thy, Tg, 5-hydroxy-5-methylhydantoine, urea, DNA damaged at Gua, some abasic sites... 

An intrastrand cross-link formed by cisplatin and adjacent guanine residues causes an unusally distorted base pair (bp) step, known as the Lippard bp step. A study of the effects of neighbouring nucleotides to the cross-linked G G has shown that the 3 -nucleotide has little effect, but the 5 -nucleotide has a dramatic effect. The 5 residue always maintains an S pucker, but the canting varies, depending on the substituent. Bleomycin causes two major lesions to DNA, formation of a 4 -keto abasic site and strand cleavage to yield a 3 -phosphog-lycolate and a 5 -phosphate. As a model for the 4 -keto abasic site, an NMR structure of a 13-mer DNA duplex containing an abasic site has been reported." It was found that for both the a- and p-anomers, the abasic site was extrahelical, and that the duplex showed very little distortion to the backbone. This was discussed in terms of repair of such lesions in vivo. 

Other modifications include a formamide residue (191), a ring fragmentation product of thymine, which exists as either a cis or a trans conformer. Both isomers are rotated out of the helix, and the bases on either side of (191) occupy the space vacated by it. A pyrene abasic site base pair in DNA duplexes adopts the usual B-form duplex, with the pyrene residue within the duplex stacking on adjacent nucleobases. The abasic site folds back over the opposite strand to shelter the hydrophobic base from exposure to water.The photoresponsive azobenzene analogues (133, R- and 5-forms) have been incorporated into DNA for NMR analysis. Both isomers intercalate between neighbouring base pairs, and the 5-isomer exhibits more disturbance in its duplex structure which is refiected in lower Tms compared to the R-isomer. ... 

The product of deoxyribose oxidation at Cl, 5, could be observed by HPLC. However, it was noted that a minor amount of C4 hydroxylation may be observed as well at some residues in the loops. The oxidation of the C4 -H bond of some deoxyribose units in the single-stranded loops was due to an hydroxylation (formation of 4 -hydroxylated abasic site, 23), no 3 -phosphogly-colate were detected. The C4 oxidation mechanism was evidenced, on polyacrylamide gels, by the protection of NaBH4 toward piperidine sensitive cleavage at some T residues. 

Fig. 7. The two structurally distinct AP endonucleases. (A) Human APEl, in blue, bound to DNA containing a substrate abasic site, in red (Mol et aL, 2000b). (B) Endonuclease IV, in red, bound to its product DNA (Hosfield et al, 1999). The abasic moiety of the dRP group is colored blue. (G) View of the APEl/DNA complex highlighting the positions of the two cysteines involved in the redox activity of this enzyme. The cysteine residues are yellow, seen buried within the interior of the protein. (See Color Insert.)...

The apparent conflict between the in vivo and in vitro results may perhaps be explained by Revlp existing within a complex in the cell, in which dCMP insertion is inhibited at template sites other than those with abasic residues and, perhaps, some other lesions. Finally, the deoxycytidyl transferase activity, even if not essential for the bypass of abasic sites, nevertheless doubles the efficiency of rephcation past this lesion in yeast (Otsuka et al., 2002b see also Section III), perhaps explaining why the activity has been maintained throughout evolution. 

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