Cocrystal structure of an editing complex of klenow fragment with dna (3'-5' exonuclease dna polymerase protein-dna interaction x-ray crystallography/metal ion catalysis)

P. S. Freemont, J. M. Friedman, L. S. Beese, M. R. Sanderson, T. A. Steitz

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

High-resolution crystal structures of editing complexes of both duplex and single-stranded DNA bound to Escherichia coli DNA polymerase I large fragment (Klenow fragment) show four nucleotides of single-stranded DNA bound to the 3′ – 5′ exonuclease active site and extending toward the polymerase active site. Melting of the duplex DNA by the protein is stabilized by hydophobic interactions between Phe-473, Leu-361, and His-666 and the last three bases at the 3′ terminus. Two divalent metal ions interacting with the phosphodiester to be hydrolyzed are proposed to catalyze the exonuclease reaction by a mechanism that may be related to mechanisms of other enzymes that catalyze phospho-group transfer including RNA enzymes. We suggest that the editing active site competes with the polymerase active site some 30 Å away for the newly formed 3′ terminus. Since a 3′ terminal mismatched base pair favors the melting of duplex DNA, its binding and excision at the editing exonuclease site that binds single-stranded DNA is enhanced.

Original languageEnglish (US)
Title of host publicationStructural Insights into Gene Expression and Protein Synthesis
PublisherWorld Scientific Publishing Co.
Pages240-244
Number of pages5
ISBN (Electronic)9789811215865
ISBN (Print)9789811215858
StatePublished - Jan 1 2020
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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