Oxidative demethylation of 3-methylthymine and 3-methyluracil in single-stranded DNA and RNA by mouse and human FTO

Jia, Guifang; Yang, Cai-Guang; Yang, Shangdong; Jian, Xing; Yi, Chengqi; Zhou, Zhiqiang; He, Chuan

doi:10.1016/j.febslet.2008.08.019

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Volume 582, Issue 23 , Pages 3313-3319, 15 October 2008

Oxidative demethylation of 3-methylthymine and 3-methyluracil in single-stranded DNA and RNA by mouse and human FTO

Edited by Ned Mantei

Guifang Jia
- Department of Chemistry, The University of Chicago, 929 East 57th Street, Chicago, IL 60637, USA
- Department of Applied Chemistry, China Agricultural University, Beijing 100094, China
- Both authors contributed equally to this work.
- This author was supported by China Scholarship Council.
Cai-Guang Yang
- Department of Chemistry, The University of Chicago, 929 East 57th Street, Chicago, IL 60637, USA
- Both authors contributed equally to this work.
Shangdong Yang
- Department of Chemistry, The University of Chicago, 929 East 57th Street, Chicago, IL 60637, USA
Xing Jian
- Department of Chemistry, The University of Chicago, 929 East 57th Street, Chicago, IL 60637, USA
Chengqi Yi
- Department of Chemistry, The University of Chicago, 929 East 57th Street, Chicago, IL 60637, USA
Zhiqiang Zhou
- Department of Applied Chemistry, China Agricultural University, Beijing 100094, China
Chuan He
- Department of Chemistry, The University of Chicago, 929 East 57th Street, Chicago, IL 60637, USA
- Corresponding author. Fax: +1 773 702 0805.

Received 29 July 2008; received in revised form 24 August 2008; accepted 25 August 2008. published online 05 September 2008.

Abstract

The human obesity susceptibility gene, FTO, encodes a protein that is homologous to the DNA repair AlkB protein. The AlkB family proteins utilize iron(II), α-ketoglutarate (α-KG) and dioxygen to perform oxidative repair of alkylated nucleobases in DNA and RNA. We demonstrate here the oxidative demethylation of 3-methylthymine (3-meT) in single-stranded DNA (ssDNA) and 3-methyluracil (3-meU) in single-stranded RNA (ssRNA) by recombinant human FTO protein in vitro. Both human and mouse FTO proteins preferentially repair 3-meT in ssDNA over other base lesions tested. They showed negligible activities against 3-meT in double-stranded DNA (dsDNA). In addition, these two proteins can catalyze the demethylation of 3-meU in ssRNA with a slightly higher efficiency over that of 3-meT in ssDNA, suggesting that methylated RNAs are the preferred substrates for FTO.

Abbreviations: mFTO, mouse FTO, hFTO, human FTO, 3-meT, 3-methylthymine, 3-meU, 3-methyluracil, 1-meA, 1-methyladenine, 1-meG, 1-methylguanine, 3-meC, 3-methylcytosine, εA, 1,N⁶-ethenoadenine, εC, 3,N⁴-ethenocytosine, IPTG, isopropyl-β-d-thiogalactopyranoside, α-KG, α-ketoglutarate, BSA, bovine serium albumin, MES, 2-(N-morpholino)ethanesulfonic acid, EDTA, ethylene diamine tetraacetic acid, TFA, trifluoroacetic acid, HPLC, high performance liquid chromatography