oxoguanine glycosylase 1, human OR C479058
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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 asparagová 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
- Názvy látek
- 8-hydroxyguanine MeSH Prohlížeč
- DNA-glykosylasy MeSH
- guanin MeSH
- kyselina asparagová MeSH
- lysin MeSH
- mutantní proteiny MeSH
- oxoguanine glycosylase 1, human MeSH Prohlížeč
OBJECTIVES: (i) to determine the extent of oxidative stress and DNA damage and repair using a panel of selected markers in patients with type 1 and type 2 diabetes mellitus (T1DM, T2DM), (ii) to find their possible relationships with diabetes compensation and duration, and finally (iii) to test for the effect of functional polymorphisms in the 8-oxoguanin DNA glycosylase (rs1052133), catalase (rs1001179) and superoxide dismutase (rs4880) genes on respective intermediate phenotypes. METHODS: A total of 207 subjects (23 children and 44 adults with T1DM, 52 adult patients with T2DM and 88 healthy adult control subjects) were enrolled in the study. The following markers of redox state were determined in participants: erythrocyte superoxide dismutase (Ery-SOD), whole blood glutathione peroxidase (WB-GPx), erythrocyte glutathione (Ery-GSH), plasma total antioxidant capacity (P-tAOC) and plasma malondialdehyde (P-MDA). Furthermore, the extent of DNA damage and repair was ascertained using the following parameters: DNA single strand breaks (DNAssb), DNA repair capacity (DNArc) and DNA repair index (DNRI). RESULTS: Comparison of T1DM vs. T2DM patients revealed significantly higher Ery-GSH content (P < 0.0001) and significantly lower Ery-SOD activity (P = 0.0006) and P-tAOC level (P < 0.0001) in T1DM subjects. T2DM diabetics exhibited a significant increase in DNAssb (P < 0.0001) and significant decrease in both DNArc (P < 0.0001) and DNRI (P < .0001) compared with T1DM patients. Patient's age (irrespective of DM type) significantly correlated with DNAssb (r = 0.48, P < 0.0001), DNArc (r = -0.67, P < 0.0001) and DNRI (r = -0.7, P < 0.0001). Allele frequencies of all studied polymorphisms did not exhibit any significant association with the investigated parameters. CONCLUSION: We demonstrated significant age- and DM type-related changes of oxidative DNA modification and capacity for its repair in subjects with T1DM and T2DM.
- MeSH
- alely MeSH
- diabetes mellitus 1. typu krev genetika MeSH
- diabetes mellitus 2. typu krev genetika MeSH
- dítě MeSH
- DNA-glykosylasy krev genetika MeSH
- dospělí MeSH
- frekvence genu MeSH
- glutathion krev MeSH
- glutathionperoxidasa krev MeSH
- katalasa krev genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- malondialdehyd krev MeSH
- oprava DNA MeSH
- oxidační stres * MeSH
- polymorfismus genetický MeSH
- poškození DNA MeSH
- senioři MeSH
- studie případů a kontrol MeSH
- superoxiddismutasa krev genetika MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
- Názvy látek
- DNA-glykosylasy MeSH
- glutathion MeSH
- glutathionperoxidasa MeSH
- katalasa MeSH
- malondialdehyd MeSH
- oxoguanine glycosylase 1, human MeSH Prohlížeč
- superoxiddismutasa MeSH
A mechanistic pathway for cleavage of the N-glycosidic bond of 8-oxo-2'-deoxyguanosine (oxoG) catalyzed with the human 8-oxoguanine glycosylase 1 DNA repair protein (hOGG1) is proposed in this theoretical study. The reaction scheme suggests direct proton addition to the glycosidic nitrogen N9 of oxoG from the Nε-ammonium of Lys249 residue of hOGG1 that is enabled owing to the N9 pyramidal geometry. The N9-pyramidalization of oxoG is induced within hOGG1 active site. The coordination of N9 nitrogen to the Nε-ammonium of Lys249 unveiled by available crystal structures enables concerted, synchronous substitution of the N9-C1' bond by the N9-H bond. The reaction is compared with other pathways already proposed by means of calculated activation energies. The ΔG(#) energy for the newly proposed reaction mechanism calculated with the B3LYP/6-31G(d,p) method 17.0 kcal mol(-1) is significantly lower than ΔG(#) energies for other reactions employing attack of the Nε-amino group to the anomeric carbon C1' of oxoG and attack of the Nε-ammonium to the N3 nitrogen of oxoG base. Moreover, activation energy for the oxoG cleavage proceeding via N9-pyramidalization is lower than energy calculated for normal G because the electronic state of the five-membered aromatic ring of oxoG is better suited for the reaction. The modification of aromatic character introduced by oxidation to the nucleobase thus seems to be the factor that is checked by hOGG1 to achieve base-specific cleavage.
- MeSH
- 8-hydroxy-2'-deoxyguanosin MeSH
- deoxyguanosin analogy a deriváty chemie metabolismus MeSH
- DNA-glykosylasy chemie metabolismus MeSH
- dusík chemie MeSH
- glykosidy chemie MeSH
- kvantová teorie MeSH
- lidé MeSH
- molekulární modely MeSH
- oprava DNA * MeSH
- termodynamika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- 8-hydroxy-2'-deoxyguanosin MeSH
- deoxyguanosin MeSH
- DNA-glykosylasy MeSH
- dusík MeSH
- glykosidy MeSH
- oxoguanine glycosylase 1, human MeSH Prohlížeč
Oxidative stress, oxidative DNA damage and resulting mutations play a role in colorectal carcinogenesis. Impaired equilibrium between DNA damage formation, antioxidant status, and DNA repair capacity is responsible for the accumulation of genetic mutations and genomic instability. The lesion-specific DNA glycosylases, e.g., hOGG1 and MUTYH, initiate the repair of oxidative DNA damage. Hereditary syndromes (MUTYH-associated polyposis, NTHL1-associated tumor syndrome) with germline mutations causing a loss-of-function in base excision repair glycosylases, serve as straight forward evidence on the role of oxidative DNA damage and its repair. Altered or inhibited function of above glycosylases result in an accumulation of oxidative DNA damage and contribute to the adenoma-adenocarcinoma transition. Oxidative DNA damage, unless repaired, often gives rise G:C > T:A mutations in tumor suppressor genes and proto-oncogenes with subsequent occurrence of chromosomal copy-neutral loss of heterozygosity. For instance, G>T transversions in position c.34 of a KRAS gene serves as a pre-screening tool for MUTYH-associated polyposis diagnosis. Since sporadic colorectal cancer represents more complex and heterogenous disease, the situation is more complicated. In the present study we focused on the roles of base excision repair glycosylases (hOGG1, MUTYH) in colorectal cancer patients by investigating tumor and adjacent mucosa tissues. Although we found downregulation of both glycosylases and significantly lower expression of hOGG1 in tumor tissues, accompanied with G>T mutations in KRAS gene, oxidative DNA damage and its repair cannot solely explain the onset of sporadic colorectal cancer. In this respect, other factors (especially microenvironment) per se or in combination with oxidative DNA damage warrant further attention. Base excision repair characteristics determined in colorectal cancer tissues and their association with disease prognosis have been discussed as well.
- Klíčová slova
- BER glycosylases, DNA repair, Oxidative DNA damage, colorectal cancer,
- MeSH
- DNA-glykosylasy * genetika metabolismus MeSH
- familiární adenomatózní polypóza MeSH
- kolorektální nádory * patologie MeSH
- lidé MeSH
- nádorové mikroprostředí MeSH
- oprava DNA genetika MeSH
- oxidační stres genetika MeSH
- protoonkogenní proteiny p21(ras) genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- DNA-glykosylasy * MeSH
- mutY adenine glycosylase MeSH Prohlížeč
- oxoguanine glycosylase 1, human MeSH Prohlížeč
- protoonkogenní proteiny p21(ras) MeSH
Sperm motility is one of the major determinants of male fertility. Since sperm need a great deal of energy to support their fast movement by active metabolism, they are thus extremely vulnerable to oxidative damage by the reactive oxygen species (ROS) and other free radicals generated as byproducts in the electron transport chain. The present study is aimed at understanding the impact of a mitochondrial oxidizing/reducing microenvironment in the etiopathology of male infertility. We detected the mitochondrial DNA (mtDNA) 4,977 bp deletion in human sperm. We examined the gene mutation of ATP synthase 6 (ATPase6 m.T8993G) in ATP generation, the gene polymorphisms of uncoupling protein 2 (UCP2, G-866A) in the uncoupling of oxidative phosphorylation, the role of genes such as manganese superoxide dismutase (MnSOD, C47T) and catalase (CAT, C-262T) in the scavenging system in neutralizing reactive oxygen species, and the role of human 8-oxoguanine DNA glycosylase (hOGG1, C1245G) in 8-hydroxy-2'-deoxyguanosine (8-OHdG) repair. We found that the sperm with higher motility were found to have a higher mitochondrial membrane potential and mitochondrial bioenergetics. The genotype frequencies of UCP2 G-866A, MnSOD C47T, and CAT C-262T were found to be significantly different among the fertile subjects, the infertile subjects with more than 50% motility, and the infertile subjects with less than 50% motility. A higher prevalence of the mtDNA 4,977 bp deletion was found in the subjects with impaired sperm motility and fertility. Furthermore, we found that there were significant differences between the occurrences of the mtDNA 4,977 bp deletion and MnSOD (C47T) and hOGG1 (C1245G). In conclusion, the maintenance of the mitochondrial redox microenvironment and genome integrity is an important issue in sperm motility and fertility.
- MeSH
- DNA-glykosylasy genetika metabolismus MeSH
- frekvence genu MeSH
- lidé MeSH
- membránový potenciál mitochondrií účinky léků MeSH
- mitochondriální DNA genetika metabolismus MeSH
- mitochondrie genetika metabolismus MeSH
- motilita spermií fyziologie MeSH
- mužská infertilita genetika patologie MeSH
- oxidační stres účinky léků MeSH
- peroxid vodíku farmakologie MeSH
- polymorfismus genetický MeSH
- spermie metabolismus fyziologie MeSH
- superoxiddismutasa genetika metabolismus MeSH
- uncoupling protein 2 genetika metabolismus MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- DNA-glykosylasy MeSH
- mitochondriální DNA MeSH
- oxoguanine glycosylase 1, human MeSH Prohlížeč
- peroxid vodíku MeSH
- superoxiddismutasa MeSH
- UCP2 protein, human MeSH Prohlížeč
- uncoupling protein 2 MeSH
Human cancers are often associated with numerical and structural chromosomal instability. Structural chromosomal aberrations (CAs) in peripheral blood lymphocytes (PBL) arise as consequences of direct DNA damage or due to replication on a damaged DNA template. In both cases, DNA repair is critical and inter-individual differences in its capacity are probably due to corresponding genetic variations. We investigated functional variants in DNA repair genes (base and nucleotide excision repair, double-strand break repair) in relation to CAs, chromatid-type aberrations (CTAs) and chromosome-type aberrations (CSAs) in healthy individuals. Chromosomal damage was determined by conventional cytogenetic analysis. The genotyping was performed by both restriction fragment length polymorphism and TaqMan allelic discrimination assays. Multivariate logistic regression was applied for testing individual factors on CAs, CTAs and CSAs. Pair-wise genotype interactions of 11 genes were constructed for all possible pairs of single-nucleotide polymorphisms. Analysed individually, we observed significantly lower CTA frequencies in association with XPD Lys751Gln homozygous variant genotype [odds ratio (OR) 0.64, 95% confidence interval (CI) 0.48-0.85, P = 0.004; n = 1777]. A significant association of heterozygous variant genotype in RAD54L with increased CSA frequency (OR 1.96, 95% CI 1.01-4.02, P = 0.03) was determined in 282 subjects with available genotype. By addressing gene-gene interactions, we discovered 14 interactions significantly modulating CAs, 9 CTAs and 12 CSAs frequencies. Highly significant interactions included always pairs from two different pathways. Although individual variants in genes encoding DNA repair proteins modulate CAs only modestly, several gene-gene interactions in DNA repair genes evinced either enhanced or decreased CA frequencies suggesting that CAs accumulation requires complex interplay between different DNA repair pathways.
- MeSH
- chromozomální aberace * MeSH
- DNA vazebné proteiny genetika MeSH
- DNA-glykosylasy genetika MeSH
- dospělí MeSH
- frekvence genu MeSH
- genetické asociační studie MeSH
- jednonukleotidový polymorfismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- nádory genetika MeSH
- oprava DNA genetika MeSH
- protein XRCC1 MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- DNA vazebné proteiny MeSH
- DNA-glykosylasy MeSH
- oxoguanine glycosylase 1, human MeSH Prohlížeč
- protein XRCC1 MeSH
Decreased levels of single-strand breaks in DNA (SSBs), reflecting DNA damage, have previously been observed with increased styrene exposure in contrast to a dose-dependent increase in the base-excision repair capacity. To clarify further the above aspects, we have investigated the associations between SSBs, micronuclei, DNA repair capacity and mRNA expression in XRCC1, hOGG1 and XPC genes on 71 styrene-exposed and 51 control individuals. Styrene concentrations at workplace and in blood characterized occupational exposure. The workers were divided into low (below 50 mg/m³) and high (above 50 mg/m³)) styrene exposure groups. DNA damage and DNA repair capacity were analyzed in peripheral blood lymphocytes by Comet assay. The mRNA expression levels were determined by qPCR. A significant negative correlation was observed between SSBs and styrene concentration at workplace (R=-0.38, p=0.001); SSBs were also significantly higher in men (p=0.001). The capacity to repair irradiation-induced DNA damage was the highest in the low exposure group (1.34±1.00 SSB/10⁹ Da), followed by high exposure group (0.72±0.81 SSB/10⁹ Da) and controls (0.65±0.82 SSB/10⁹ Da). The mRNA expression levels of XRCC1, hOGG1 and XPC negatively correlated with styrene concentrations in blood and at workplace (p<0.001) and positively with SSBs (p<0.001). Micronuclei were not affected by styrene exposure, but were higher in older persons and in women (p<0.001). In this study, we did not confirm previous findings on an increased DNA repair response to styrene-induced genotoxicity. However, negative correlations of SSBs and mRNA expression levels of XRCC1, hOGG1 and XPC with styrene exposure warrant further highly-targeted study.
- MeSH
- DNA vazebné proteiny biosyntéza genetika MeSH
- DNA-glykosylasy biosyntéza genetika MeSH
- dospělí MeSH
- jednořetězcové zlomy DNA účinky léků MeSH
- kometový test MeSH
- lidé středního věku MeSH
- lidé MeSH
- lymfocyty účinky léků MeSH
- messenger RNA biosyntéza MeSH
- mladý dospělý MeSH
- oprava DNA účinky léků genetika MeSH
- pracovní expozice škodlivé účinky MeSH
- protein XRCC1 MeSH
- regulace genové exprese účinky léků MeSH
- styren škodlivé účinky krev MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH
- Názvy látek
- DNA vazebné proteiny MeSH
- DNA-glykosylasy MeSH
- messenger RNA MeSH
- oxoguanine glycosylase 1, human MeSH Prohlížeč
- protein XRCC1 MeSH
- styren MeSH
- XPC protein, human MeSH Prohlížeč
- XRCC1 protein, human MeSH Prohlížeč
DNA integrity was investigated in the lymphocytes of 50 bus drivers, 20 garagemen and 50 controls using the comet assay with excision repair enzymes. In parallel, 8-oxo-7,8-dihydro-2'-deoxyguanosine and 15-F(2t)-isoprostane levels in the urine and protein carbonyl levels in the plasma were assessed as markers of oxidative damage to DNA, lipids and proteins. Exposure to carcinogenic polycyclic aromatic hydrocarbons (cPAHs) and volatile compounds was measured by personal samplers for 48 and 24h, respectively, before the collection of biological specimens. Both exposed groups exhibited a higher levels of DNA instability and oxidative damage to biological macromolecules than the controls. The incidence of oxidized lesions in lymphocyte DNA, but not the urinary levels of 8-oxodG, correlated with exposure to benzene and triglycerides increased this damage. Oxidative damage to lipids and proteins was associated with exposure to cPAHs and the lipid peroxidation levels positively correlated with age and LDL cholesterol, and negatively with vitamin C. The carriers of at least one variant hOGG1 (Cys) allele tended to higher oxidative damage to lymphocyte DNA than those with the wild genotype, while XPD23 (Gln/Gln) homozygotes were more susceptible to the induction of DNA strand breaks. In contrast, GSTM1 null variant seemed to protect DNA integrity.
- MeSH
- 8-hydroxy-2'-deoxyguanosin analogy a deriváty MeSH
- dinoprost analogy a deriváty moč MeSH
- DNA-glykosylasy genetika metabolismus MeSH
- DNA účinky léků MeSH
- genetická predispozice k nemoci MeSH
- glutathiontransferasa genetika MeSH
- guanin analogy a deriváty moč MeSH
- karbonylace proteinů účinky léků MeSH
- kometový test MeSH
- látky znečišťující vzduch v pracovním prostředí toxicita MeSH
- lidé MeSH
- lymfocyty chemie účinky léků MeSH
- oxidační stres účinky léků MeSH
- peroxidace lipidů účinky léků MeSH
- polycyklické aromatické uhlovodíky analýza toxicita MeSH
- polymorfismus genetický * MeSH
- poškození DNA MeSH
- těkavé organické sloučeniny analýza toxicita MeSH
- vazokonstriktory moč MeSH
- výfukové emise vozidel toxicita MeSH
- xeroderma pigmentosum - protein skupiny D genetika MeSH
- znečištění ovzduší * MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- 8-epi-prostaglandin F2alpha MeSH Prohlížeč
- 8-hydroxy-2'-deoxyguanosin MeSH
- 8-oxo-7,8-dihydrodeoxyguanine MeSH Prohlížeč
- dinoprost MeSH
- DNA-glykosylasy MeSH
- DNA MeSH
- ERCC2 protein, human MeSH Prohlížeč
- glutathione S-transferase M1 MeSH Prohlížeč
- glutathiontransferasa MeSH
- guanin MeSH
- látky znečišťující vzduch v pracovním prostředí MeSH
- oxoguanine glycosylase 1, human MeSH Prohlížeč
- polycyklické aromatické uhlovodíky MeSH
- těkavé organické sloučeniny MeSH
- vazokonstriktory MeSH
- výfukové emise vozidel MeSH
- xeroderma pigmentosum - protein skupiny D MeSH
The (15)N NMR shifts of 9-ethyl-8-oxoguanine (OG) were calculated and measured in liquid DMSO and in crystal. The OG molecule is a model for oxidatively damaged 2'-deoxyguanosine that occurs owing to oxidative stress in cell. The DNA lesion is repaired with human 8-oxoguanine glycosylase 1 (hOGG1) base-excision repair enzyme, however, the exact mechanism of excision of damaged nucleobase with hOGG1 is currently unknown. This benchmark study on (15)N NMR shifts of OG aims their accurate structural interpretation and calibration of the calculation protocol utilizable in future studies on mechanism of hOGG1 enzyme. The effects of NMR reference, DFT functional, basis set, solvent, structure, and dynamics on calculated (15)N NMR shifts were first evaluated for OG in crystal to calibrate the best performing calculation method. The effect of large-amplitude motions on (15)N NMR shifts of OG in liquid was calculated employing molecular dynamics. The B3LYP method with Iglo-III basis used for B3LYP optimized geometry with 6-311++G(d,p) basis and including effects of solvent and molecular dynamic was the calculation protocol used for calculation of (15)N NMR shifts of OG. The NMR shift of N9 nitrogen of OG was particularly studied because the atom is involved in an N-glycosidic bond that is cleaved with hOGG1. The change of N9 NMR shift owing to oxidation of 9-ethylguanine (G) measured in liquid was -27.1 ppm. The calculated N9 NMR shift of OG deviated from experiment in crystal and in liquid by 0.45 and 0.65 ppm, respectively. The calculated change of N9 NMR shift owing to notable N9-pyramidalization of OG in one previously found polymorph was 20.53 ppm. We therefore assume that the pyramidal geometry of N9 nitrogen that could occur for damaged DNA within hOGG1 catalytic site might be detectable with (15)N NMR spectroscopy. The calculation protocol can be used for accurate structural interpretation of (15)N NMR shifts of oxidatively damaged guanine DNA residue.
