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PMID: 15494448 Published · epublish English Journal Article

The C-terminal alphaO helix of human Ogg1 is essential for 8-oxoguanine DNA glycosylase activity: the mitochondrial beta-Ogg1 lacks this domain and does not have glycosylase activity.

Nucleic acids research ·Vol. 32 ·No. 18 ·2004-00-00 ·Pages 5596-608

Hashiguchi K, Stuart JA, de Souza-Pinto NC, Bohr VA

Abstract

The human Ogg1 glycosylase is responsible for repairing 8-oxo-7,8-dihydroguanine (8-oxoG) in both nuclear and mitochondrial DNA. Two distinct Ogg1 isoforms are present; alpha-Ogg1, which mainly localizes to the nucleus and beta-Ogg1, which localizes only to mitochondria. We recently showed that mitochondria from rho(0) cells, which lack mitochondrial DNA, have similar 8-oxoG DNA glycosylase activity to that of wild-type cells. Here, we show that beta-Ogg1 protein levels are approximately 80% reduced in rho(0) cells, suggesting beta-Ogg1 is not responsible for 8-oxoG incision in mitochondria. Thus, we characterized the biochemical properties of recombinant beta-Ogg1. Surprisingly, recombinant beta-Ogg1 did not show any significant 8-oxoG DNA glycosylase activity in vitro. Since beta-Ogg1 lacks the C-terminal alphaO helix present in alpha-Ogg1, we generated mutant proteins with various amino acid substitutions in this domain. Of the seven amino acid positions substituted (317-323), we identified Val-317 as a novel critical residue for 8-oxoG binding and incision. Our results suggest that the alphaO helix is absolutely necessary for 8-oxoG DNA glycosylase activity, and thus its absence may explain why beta-Ogg1 does not catalyze 8-oxoG incision in vitro. Western blot analysis revealed the presence of significant amounts of alpha-Ogg1 in human mitochondria. Together with previous localization studies in vivo, this suggests that alpha-Ogg1 protein may provide the 8-oxoG DNA glycosylase activity for the repair of these lesions in human mitochondrial DNA. beta-Ogg1 may play a novel role in human mitochondria.

MeSH Terms
Amino Acid Sequence DNA Glycosylases/chemistry,metabolism Guanine/analogs & derivatives,metabolism Humans Mitochondria/enzymology Mitochondrial Proteins/chemistry,metabolism Molecular Sequence Data Phenylalanine/chemistry Protein Structure, Secondary Protein Structure, Tertiary
Chemicals
Mitochondrial Proteins Phenylalanine 8-hydroxyguanine Guanine DNA Glycosylases oxoguanine glycosylase 1, human
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Hashiguchi K
Laboratory of Molecular Gerontology, National Institute of Aging-IRP, National Institutes of Health, Box1, 5600 Nathan Shock Drive, Baltimore, MD 21224, USA.
Stuart J A
de Souza-Pinto N C
Bohr V A
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
1362-4962
DOI
10.1093/nar/gkh863
Published
2004-00-00
Epub
2004-00-19
Pages
5596-608
Language
English
Region
England
NLM ID
0411011
PMCID
PMC524278
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/gkh863 PMC Full Text View journal details
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