The anatomy of infidelity

© 2001 Nature Publishing Group http://structbio.nature.com

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The anatomy of infidelity
Tom Ellenberger and Laura F. Silvian
Recent crystal structures of error prone DNA polymerases that bypass damage in DNA templates provide counterexamples to high fidelity polymerases.

© 2001 Nature Publishing Group http://structbio.nature.com

The number of DNA polymerases skyrocketedrecently with the discovery of a very large group of error prone DNA polymerases, the Y-superfamily1, capable of waltzing past DNA lesions that trip up normal replicative polymerases. Structures of two of these remarkable enzymes were recently reported. The crystal structures of the Saccharomyces cerevisiae RAD30A polymerase (Pol η)2 and the Sulfolobus solfataricus Dbh polymerase3 provide theinitial glimpses of Y-superfamily polymerases that restart replication at sites of mismatched or damaged bases4–9 and that contribute to adaptive mutagenesis in cells growing under adverse conditions10,11. Error prone polymerases allow cells to cope with unrepaired DNA damage by enabling the completion of replication in the face of otherwise terminal roadblocks. These ‘sloppier copiers’ lack many ofthe virtues of other polymerases, including speed, fidelity and a firm grip (processivity) on the DNA template. However, the biological fitness of the error prone DNA polymerases should be judged by a different standard. They are, after all, specialists in resolving messy situations that make respectable polymerases blush. It was unforeseen that cells would be equipped with such a vast number ofpolymerases to deal with replication mistakes and chemical insults to DNA. The sheer number of lesion bypass polymerases, and in particular the existence of many orthologs in higher organisms, suggests that individual enzymes have highly specialized cellular roles. Additional support for this proposal comes from the unique biochemical characteristics of various Y-superfamily polymerases, includingtheir different efficiencies in bypassing particular lesions and their mutagenic propensities12–18. The broad phylogenetic distribution of the lesion bypass polymerases underscores the strategic importance of tolerating DNA damage by replication bypass as a means of survival when DNA repair cannot be completed in a timely manner. Although we do not fully understand the molecular logic behind thedecision to either repair a DNA lesion

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b

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Fig. 1 Comparison of error prone and high fidelity polymerases. There is no sequence similarity between the error prone DNA polymerases, represented here by Sulfolobus solfataricus Dbh3 and Saccharomyces cerevisiae Pol η2, and highly accurate replicative polymerases like the bacteriophage T7 DNA polymerase31. Nonetheless, all three polymeraseshave similar shapes, resembling a right hand composed of fingers (yellow), palm (blue) and thumb (red) subdomains. Dbh and Pol η have finger and thumb domains that are smaller than those of T7 DNA polymerase. This creates a shallow and unencumbered active site in the error prone polymerases. The surfaces of all three enzymes are shown from the vantage point of DNA exiting the polymerase. a, TheS. solfataricus Dbh catalytic fragment (residues 1–205, PDB accession 1IM4). b, The S. cerevisiae pol η polymerase fragment (residues 1–509, PDB accession 1JIH) contains an additional C-terminal polymerase associated domain (green) that is proposed to contact DNA. c, The T7-DNA polymerase (PDB accession 1T7P) is composed of palm, fingers and thumb subdomains with additional proofreading exonuclease(gray) and processivity domains (tan). The extended thumb of T7 DNA polymerase and its more expansive fingers create an enclosed substrate binding site that undoubtably contributes to the high fidelity of this polymerase. The DNA and nucleotide substrates have been omitted from the T7 structure in order to show the active site.

or replicate past it, both options appear to be important. Human...