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Inhibition of O6-methylguanine-DNA methyltransferase by an alkyltransferase-like protein from Escherichia coli

Lookup NU author(s): Dr Mauro Santibanez Koref



The alkyltransferase-like (ATL) proteins contain primary sequence motifs resembling those found in DNA repair O6-alkylguanine-DNA alkyltransferase proteins. However, in the putative active site of ATL proteins, a tryptophan (W83) residue replaces the cysteine at the known active site of alkyltransferases. The Escherichia coli atl gene was expressed as a fusion protein and purified. Neither ATL nor C83 or A83 mutants transferred [3H] from [3H]-methylated DNA to themselves, and the levels of O6-methyl guanine (O6-meG) in substrate DNA were not affected by ATL. However, ATL inhibited the transfer of methyl groups to human alkyltransferase (MGMT). Inhibition was reduced by prolonged incubation in the presence of MGMT, again suggesting that O6-meG in the substrate is not changed by ATL. Gel-shift assays show that ATL binds to short single- or double-stranded oligonucleotides containing O6-meG, but not to oligonucleotides containing 8-oxoguanine, ethenoadenine, 5-hydroxycytosine or O4-methylthymine. There was no evidence of demethylation of O6-meG or of glycosylase or endonuclease activity. Overexpression of ATL in E.coli increased, or did not affect, the toxicity of N-methyl-N′-nitro-N-nitrosoguanidine in an alklyltransferase-proficient and -deficient strain, respectively. These results suggest that ATL may act as a damage sensor that flags O6-meG and possibly other O6-alkylation lesions for processing by other repair pathways. © The Author 2005. Published by Oxford University Press. All rights reserved.

Publication metadata

Author(s): Pearson SJ, Ferguson J, Santibanez-Koref M, Margison GP

Publication type: Article

Publication status: Published

Journal: Nucleic Acids Research

Year: 2005

Volume: 33

Issue: 12

Pages: 3837-3844

Print publication date: 01/01/2005

Date deposited: 31/01/2011

ISSN (print): 0305-1048

ISSN (electronic): 1362-4962

Publisher: Oxford University Press


DOI: 10.1093/nar/gki696

PubMed id: 16027108


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