Escherichia coli single-stranded DNA binding protein SSB promotes AlkB-mediated DNA dealkylation repair

Nigam, Richa and Roy, Anindya (2018) Escherichia coli single-stranded DNA binding protein SSB promotes AlkB-mediated DNA dealkylation repair. Biochemical and Biophysical Research Communications. ISSN 0006291X (In Press)

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Repair of alkylation damage in DNA is essential for maintaining genome integrity. Escherichia coli (E.coli) protein AlkB removes various alkyl DNA adducts including N1-methyladenine (N1meA) and N3-methylcytosine (N3meC) by oxidative demethylation. Previous studies showed that AlkB preferentially removes N1meA and N3meC from single-stranded DNA (ssDNA). It can also remove N1meA and N3meC from double-stranded DNA by base-flipping. Notably, ssDNA produced during DNA replication and recombination, remains bound to E. coli single-stranded DNA binding protein SSB and it is not known whether AlkB can repair methyl adduct present in SSB-coated DNA. Here we have studied AlkB-mediated DNA repair using SSB-bound DNA as substrate. In vitro repair reaction revealed that AlkB could efficiently remove N3meC adducts inasmuch as DNA length is shorter than 20 nucleotides. However, when longer N3meC-containing oligonuleotides were used as the substrate, efficiency of AlkB catalyzed reaction was abated compared to SSB-bound DNA substrate of identical length. Truncated SSB containing only the DNA binding domain could also support the stimulation of AlkB activity, suggesting the importance of SSB-DNA interaction for AlkB function. Using 70-mer oligonucleotide containing single N3meC we demonstrate that SSB-AlkB interaction promotes faster repair of the methyl DNA adducts.

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IITH Creators:
IITH CreatorsORCiD
Roy, Anindya
Item Type: Article
Uncontrolled Keywords: Fe(II)/2-oxoglutarate-dependent dioxygenase, AlkB, SSB, DNA repair
Subjects: Others > Biotechnology
Divisions: Department of Biotechnology
Depositing User: Team Library
Date Deposited: 16 Jan 2018 10:10
Last Modified: 16 Jan 2018 10:10
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