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• W1985331505 endingPage "10757" @default.
• W1985331505 startingPage "10748" @default.
• W1985331505 abstract "DNA polymerase selectivity is crucial for the survival of any living species, yet varies significantly among different DNA polymerases. Errors within DNA polymerase-catalyzed DNA synthesis result from the insertion of noncanonical nucleotides and extension of misaligned DNA substrates. The substrate binding characteristics among DNA polymerases are believed to vary in properties such as shape and tightness of the binding pocket, which might account for the observed differences in fidelity. Here, we employed 4′-alkylated nucleotides and primer strands bearing 4′-alkylated nucleotides at the 3′-terminal position as steric probes to investigate differential active site properties of human DNA polymerase β (Pol β) and the 3′→5′-exonuclease-deficient Klenow fragment of E. coli DNA polymerase I (KF(exo-)). Transient kinetic measurements indicate that both enzymes vary significantly in active site tightness at both positions. While small 4′-methyl and -ethyl modifications of the nucleoside triphosphate perturb Pol β catalysis, extension of modified primer strands is only marginally affected. Just the opposite was observed for KF(exo-). Here, incorporation of the modified nucleotides is only slightly reduced, whereas size augmentation of the 3′-terminal nucleotide in the primer reduces the catalytic efficiency by more than 7000- and 260 000-fold, respectively. NMR studies support the notion that the observed effects derive from enzyme substrate interactions rather than inherent properties of the modified substrates. These findings are consistent with the observed differential capability of the investigated DNA polymerases in fidelity such as processing misaligned DNA substrates. The results presented provide direct evidence for the involvement of varied steric effects among different DNA polymerases on their fidelity." @default.
• W1985331505 created "2016-06-24" @default.
• W1985331505 creator A5017489146 @default.
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• W1985331505 creator A5082422877 @default.
• W1985331505 creator A5090583758 @default.
• W1985331505 date "2008-07-16" @default.
• W1985331505 modified "2023-10-15" @default.
• W1985331505 title "Opposed Steric Constraints in Human DNA Polymerase β and <i>E. coli</i> DNA Polymerase I" @default.
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• W1985331505 doi "https://doi.org/10.1021/ja8028284" @default.
• W1985331505 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/18627154" @default.
• W1985331505 hasPublicationYear "2008" @default.
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