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The m6A methylation perturbs Hoogsteen pairing-guided incorporation of an oxidized nucleotide

Abstract

Natural nucleic acid bases can form Watson-Crick (WC) or Hoogsteen (HG) base pairs. Importantly, 8-oxo-2’-deoxyguanosine (8-oxo-dG) in DNA or 8-oxo-dG 5’-triphosphate (8-oxo-dGTP) favors a syn conformation because of the steric repulsion between O8 and O4’ of the deoxyribose ring. 8-oxo-dGTP can be incorporated into DNA opposite the templating adenine (A) using HG pairing as the dominant mechanism. Both RNA and DNA can be methylated at the N6 position of A to form N6-methyladenine (m6A). It has been found that certain viral infection may trigger an increase in production of both 8-oxo-dGTP and m6A. The current study aims to systematically explore the effects of m6A methylation on HG base pairs and the consequent nucleotide incorporation. Our thermodynamic melting study shows that the m6A·8-oxo-dG is significantly less stable than the A·8-oxo-dG base pair in paired region of DNA duplex. Moreover, we have used pre-steady-state kinetics to examine the incorporation of 8-oxo-dGTP opposite m6A relative to A by a variety of reverse transcriptase (RT) enzymes and DNA polymerase (DNA pol) enzymes, such as human immunodeficiency virus type 1 (HIV-1) RT and human DNA pol β. The results demonstrate that all these enzymes incorporate 8-oxo-dGTP less efficiently opposite m6A relative to A. Considering the steric bulk of the purine-purine pair between 8-oxo-dG and A, m6A methylation may affect the HG pairing to a great extent. Hence, it will be unfavorable to incorporate 8-oxo-dGTP into the growing strand opposite m6A. Moreover, the impeded incorporation of 8-oxo-dGTP opposite m6A has been extended to determine m6A at pre-defined positions in human rRNA. Our study may provide new insights into roles of m6A in reducing the mutagenic potential of cellular 8-oxo-dGTP.

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Publication details

The article was received on 24 May 2017, accepted on 05 Jul 2017 and first published on 06 Jul 2017


Article type: Edge Article
DOI: 10.1039/C7SC02340E
Citation: Chem. Sci., 2017, Accepted Manuscript
  • Open access: Creative Commons BY license
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    The m6A methylation perturbs Hoogsteen pairing-guided incorporation of an oxidized nucleotide

    X. Zhou, S. Wang, Y. Song, Y. Wang, X. Li, B. Fu, Y. Liu, J. Wang, L. Wei and T. Tian, Chem. Sci., 2017, Accepted Manuscript , DOI: 10.1039/C7SC02340E

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