DNA structure and polymerase fidelity

The accuracy of DNA replication results from both the intrinsic DNA polymer ase fidelity and the DNA sequence. Although the recent structural studies o n polymerases have brought new insights on polymerase fidelity, the role of DNA sequence and structure is less well understood. Here, the analysis of the crystal structures of hotspots for polymerase slippage including (CA), and (A), tracts in different intermolecular con texts reveals that, in the B-form, these sequences share common structural alterations which may expla in the high rate of replication errors. In particular, a two-faced "Janus-l ike" structure with shifted base-pairs in the major groove but an apparent normal geometry in the minor groove constitutes a molecular decoy specifica lly suitable to mislead the polymerases. A model of the rat polymerase beta bound to this structure suggests that an altered conformation of the nasce nt template-primer duplex can interfere with correct nucleotide incorporati on by affecting the geometry of the active site and breaking the rules of b ase-pairing, while at the same time escaping enzymatic mechanisms of error discrimination which scan for the correct geometry of the minor groove. In contrast, by showing that the A-form greatly attenuates the sequence-dep endent structural alterations in hotspots, this study suggests that the A-c onformation of the nascent template-primer duplex at the vicinity of the po lymerase active site will contribute to fidelity. The A-form may play the r ole of a structural buffer which preserves the correct geometry of the acti ve site for all sequences. The detailed comparison of the conformation of t he nascent template-primer duplex in the available crystal structures of DN A polymerase-DNA complexes shows that polymerase beta, the least accurate e nzyme, is unique in binding to a B-DNA duplex even close to its active site . This model leads to several predictions which are discussed in the light of published experimental data. (C) 1999 Academic Press.