Abstract
The ability or inability of a DNA polymerase to extend a mispair directly affects the establishment of genomic mutations. We report here kinetic analyses of the ability of Dpo4, a Y-family polymerase from Sulfolobus solfataricus, to extend from all mispairs opposite a template G or T. Dpo4 is equally inefficient at extending these mispairs, which include, surprisingly, a G·T mispair expected to conform closely to Watson-Crick geometry. To elucidate the basis of this, we solved the structure of Dpo4 bound to G·T-mispaired primer template in the presence of an incoming nucleotide. As a control, we also determined the structure of Dpo4 bound to a matched A-T base pair at the primer terminus. The structures offer a basis for the low efficiency of Dpo4 in extending a G·T mispair: a reverse wobble that deflects the primer 3′-OH away from the incoming nucleotide.
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Acknowledgements
We thank the staff at the Advanced Photon Source and the National Synchrotron Light Source for facilitating X-ray data collection. We thank T. Edwards for help with data collection and processing. This was supported was supported by US National Institutes of Health grant CA094006 (A.K.A. and L.P.).
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Trincao, J., Johnson, R., Wolfle, W. et al. Dpo4 is hindered in extending a G·T mismatch by a reverse wobble. Nat Struct Mol Biol 11, 457–462 (2004). https://doi.org/10.1038/nsmb755
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DOI: https://doi.org/10.1038/nsmb755
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