7r3x
The crystal structure of the L439V variant of Pol2CORE in complex with DNA and an incoming nucleotide
Structural highlights
FunctionDPOE_YEAST DNA polymerase epsilon (DNA polymerase II) participates in chromosomal DNA replication. It is required during synthesis of the leading and lagging DNA strands at the replication fork and binds at/or near replication origins and moves along DNA with the replication fork. It has 3'-5' proofreading exonuclease activity that correct errors arising during DNA replication. It is also involved in DNA synthesis during DNA repair.[1] Publication Abstract from PubMedAmino acid substitutions in the exonuclease domain of DNA polymerase (Pol) cause ultramutated tumors. Studies in model organisms suggested pathogenic mechanisms distinct from a simple loss of exonuclease. These mechanisms remain unclear for most recurrent Pol mutations. Particularly, the highly prevalent V411L variant remained a long-standing puzzle with no detectable mutator effect in yeast despite the unequivocal association with ultramutation in cancers. Using purified four-subunit yeast Pol, we assessed the consequences of substitutions mimicking human V411L, S459F, F367S, L424V and D275V. While the effects on exonuclease activity vary widely, all common cancer-associated variants have increased DNA polymerase activity. Notably, the analog of Pol-V411L is among the strongest polymerases, and structural analysis suggests defective polymerase-to-exonuclease site switching. We further show that the V411L analog produces a robust mutator phenotype in strains that lack mismatch repair, indicating a high rate of replication errors. Lastly, unlike wild-type and exonuclease-dead Pol, hyperactive variants efficiently synthesize DNA at low dNTP concentrations. We propose that this characteristic could promote cancer cell survival and preferential participation of mutator polymerases in replication during metabolic stress. Our results support the notion that polymerase fitness, rather than low fidelity alone, is an important determinant of variant pathogenicity. Enhanced polymerase activity permits efficient synthesis by cancer-associated DNA polymerase variants at low dNTP levels.,Barbari SR, Beach AK, Markgren JG, Parkash V, Moore EA, Johansson E, Shcherbakova PV Nucleic Acids Res. 2022 Aug 12;50(14):8023-8040. doi: 10.1093/nar/gkac602. PMID:35822874[2] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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