DNA binding mechanism of O6-alkylguanine-DNA alkyltransferase: stoichiometry and effects of DNA base composition and secondary structure on complex stability

Biochemistry
Michael G FriedA E Pegg

Abstract

O6-Alkylguanine-DNA alkyltransferase (AGT) is an important cellular defense against the mutagenic effects of DNA alkylating agents. In humans this defense can contribute to the ability of some tumors to resist the effects of chemotherapeutic agents that act through DNA alkylation. We report here studies that characterize the interaction of AGT with DNA. We show that although AGT sediments as a monomer in the absence of DNA, it binds cooperatively to single stranded deoxyribonucleotides. The stoichiometries of complexes formed with 16-, 30-, and 80-base oligodeoxyribonucleotides are 3.8 +/- 0.3, 5.3 +/- 0.2, and 8.9 +/- 0.2, respectively; the binding density decreasing from approximately 4 nt/monomer to approximately 9 nt/monomer as DNA length increases over this range. Binding competition assays show that DNA affinities depend only weakly on base composition or secondary structure, although in general G + C-rich sequences are bound with greater affinity than are A + T-rich ones and single-stranded DNA is bound with greater affinity than duplex forms. These results suggest mechanisms by which AGT may search for alkylated sites and interact with them to effect DNA repair.

References

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Citations

Jun 18, 2009·The Journal of Biological Chemistry·Rebecca GuzaNatalia Tretyakova
Dec 23, 2009·The Journal of Biological Chemistry·Qingming FangAnthony E Pegg
Dec 2, 2006·The Journal of Biological Chemistry·Joseph J RasimasMichael G Fried
Sep 23, 2000·Nucleic Acids Research·P E VerdematoL Y Lian
Dec 22, 1999·Nucleic Acids Research·J E WibleyP C Moody
Aug 2, 2012·Organic & Biomolecular Chemistry·Francis P McManusChristopher J Wilds
Jul 11, 2006·Magnetic Resonance in Chemistry : MRC·Anne RobertsDavid E Wemmer
Aug 5, 2014·Chembiochem : a European Journal of Chemical Biology·Francis P McManus, Christopher J Wilds
May 16, 2006·Cancer Treatment Reviews·Cara A RabikM Eileen Dolan
May 29, 2015·Biopolymers·Manana Melikishvili, Michael G Fried
Apr 11, 2009·Journal of Molecular Biology·Claire A AdamsMichael G Fried
Sep 25, 2017·Organic & Biomolecular Chemistry·Alexey Y DenisovChristopher J Wilds
Nov 16, 2018·Organic & Biomolecular Chemistry·William CoppChristopher J Wilds
Dec 24, 2002·The Journal of Biological Chemistry·Joseph J RasimasMichael G Fried
Jun 3, 2017·Nucleic Acids Research·Manana Melikishvili, Michael G Fried

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