The excision of 8-oxoguanine (oxoG) by the human 8-oxoguanine DNA glycosylase 1 (hOGG1) base-excision repair enzyme was studied by using the QM/MM (M06-2X/6-31G(d,p):OPLS2005) calculation method and nuclear magnetic resonance (NMR) spectroscopy. The calculated glycosylase reaction included excision of the oxoG base, formation of Lys249-ribose enzyme-substrate covalent adduct and formation of a Schiff base. The formation of a Schiff base with ΔG# = 17.7 kcal/mol was the rate-limiting step of the reaction. The excision of the oxoG base with ΔG# = 16.1 kcal/mol proceeded via substitution of the C1΄-N9 N-glycosidic bond with an H-N9 bond where the negative charge on the oxoG base and the positive charge on the ribose were compensated in a concerted manner by NH3+(Lys249) and CO2-(Asp268), respectively. The effect of Asp268 on the oxoG excision was demonstrated with 1H NMR for WT hOGG1 and the hOGG1(D268N) mutant: the excision of oxoG was notably suppressed when Asp268 was mutated to Asn. The loss of the base-excision function was rationalized with QM/MM calculations and Asp268 was confirmed as the electrostatic stabilizer of ribose oxocarbenium through the initial base-excision step of DNA repair. The NMR experiments and QM/MM calculations consistently illustrated the base-excision reaction operated by hOGG1.
- MeSH
- biokatalýza MeSH
- DNA-glykosylasy chemie metabolismus MeSH
- guanin analogy a deriváty metabolismus MeSH
- kyselina aspartová metabolismus MeSH
- lidé MeSH
- lysin metabolismus MeSH
- molekulární modely MeSH
- mutantní proteiny chemie metabolismus MeSH
- oprava DNA * MeSH
- protonová magnetická rezonanční spektroskopie MeSH
- termodynamika MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
This article suggests a new mechanistic scheme of the catalytic 8-oxoguanine excision with the hOGG1 base excision repair protein. The energy-efficient and substratespecific scheme employs enforced pyramidalization of the glycosidic nitrogen in the nucleobase within the hOGG1 catalytic pocket.
- Klíčová slova
- human 8-oxoguanine glycosylase1 protein,
- MeSH
- chemické jevy MeSH
- deoxyguanosin chemie MeSH
- DNA-glykosylasy * chemie MeSH
- enzymy opravy DNA * chemie MeSH
- lidé MeSH
- molekulární modely MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
We have determined the three-dimensional (3D) structure of DNA duplex that includes tandem Hg(II)-mediated T-T base pairs (thymine-Hg(II)-thymine, T-Hg(II)-T) with NMR spectroscopy in solution. This is the first 3D structure of metallo-DNA (covalently metallated DNA) composed exclusively of 'NATURAL' bases. The T-Hg(II)-T base pairs whose chemical structure was determined with the (15)N NMR spectroscopy were well accommodated in a B-form double helix, mimicking normal Watson-Crick base pairs. The Hg atoms aligned along DNA helical axis were shielded from the bulk water. The complete dehydration of Hg atoms inside DNA explained the positive reaction entropy (ΔS) for the T-Hg(II)-T base pair formation. The positive ΔS value arises owing to the Hg(II) dehydration, which was approved with the 3D structure. The 3D structure explained extraordinary affinity of thymine towards Hg(II) and revealed arrangement of T-Hg(II)-T base pairs in metallo-DNA.
