Title: Mechanism of error-free DNA synthesis across N1-methyl-deoxyadenosine by human DNA polymerase-ι.

Authors: Jain, Rinku; Choudhury, Jayati Roy; Buku, Angeliki; Johnson, Robert E; Prakash, Louise; Prakash, Satya; Aggarwal, Aneel K

Published In Sci Rep, (2017 Mar 08)

Abstract: N1-methyl-deoxyadenosine (1-MeA) is formed by methylation of deoxyadenosine at the N1 atom. 1-MeA presents a block to replicative DNA polymerases due to its inability to participate in Watson-Crick (W-C) base pairing. Here we determine how human DNA polymerase-ι (Polι) promotes error-free replication across 1-MeA. Steady state kinetic analyses indicate that Polι is ~100 fold more efficient in incorporating the correct nucleotide T versus the incorrect nucleotide C opposite 1-MeA. To understand the basis of this selectivity, we determined ternary structures of Polι bound to template 1-MeA and incoming dTTP or dCTP. In both structures, template 1-MeA rotates to the syn conformation but pairs differently with dTTP versus dCTP. Thus, whereas dTTP partakes in stable Hoogsteen base pairing with 1-MeA, dCTP fails to gain a "foothold" and is largely disordered. Together, our kinetic and structural studies show how Polι maintains discrimination between correct and incorrect incoming nucleotide opposite 1-MeA in preserving genome integrity.

PubMed ID: 28272441

MeSH Terms: Base Pairing; Catalytic Domain; Crystallography, X-Ray; DNA Polymerase iota; DNA-Directed DNA Polymerase/chemistry; DNA-Directed DNA Polymerase/metabolism*; DNA/biosynthesis*; DNA/chemistry; Deoxyadenosines/chemistry; Deoxyadenosines/metabolism*; Deoxycytosine Nucleotides/chemistry; Deoxycytosine Nucleotides/metabolism; Humans; Kinetics; Protein Structure, Quaternary; Thymine Nucleotides/chemistry; Thymine Nucleotides/metabolism