Analysis of Novel Intermediate of Guanine-Guanine Crosslink Produced in Reactions of One-Electron Oxidation of Guanine Derivatives by Using 8-Substituted 2´-Deoxyguanosines as Analog Compounds

Oxidative damage to DNA has been implicated in a plethora of pathologies, such as cancer, neurodegenerative diseases, cardiovascular diseases, and aging. One-electron transfer (OET) plays a significant role in oxidative DNA damage in vivo. Guanine as the most oxidizable part of DNA is the major focus of studies on oxidation damage to DNA initiated by OET. Until recently, the pathway of guanine one-electron oxidation via its neutral guanine radical, G·, has been poorly studied. Our recent research has discovered a novel type of products of G· dimerization, D1 and D2, formed as a result of oxidation reaction of guanine derivatives, initiated by OET. A proposed reaction mechanism contains an early intermediate (Int1) generated by recombination of the two G· radicals. We were not able to isolate Int1, so that its role in the proposed reaction mechanism is only hypothetical. Literature data have reported that 8-arylamino-substituted 2´-deoxyguanosine (dGuo) compounds can be oxidized to create structural analogs of D1 and D2. As a result, the original 8-substituted dGuo compounds can serve as analogs of Int1. The goal of this work is therefore to confirm that Int1 is a precursor to D1 and D2 using the analogy approach. Two 8-aryl- and three 8-alkyl-substituted dGuo compounds were synthesized, purified by semipreparative HPLC, and their structures were confirmed by 1H-NMR. 8-subsituted oxidation products analogous to D1 and D2 were obtained from 8-substituted dGuo analogs upon illumination the reaction mixture in the presence of S2O82- as a oxidant and Ru(II)bpy32+ as a photosensitizer at 470 nm. The products were purified by semipreparative HPLC, and their structures were confirmed by 1H-NMR. The purified analogs of D1 were successfully tested for conversion into the D2 analog. Finally, the analogs of D2 were successfully tested for the reaction with primary amines to form analogs of 2-aminoimidozalone (AIz), in agreement with the mechanism characteristic of D2

Binding of the human nucleotide excision repair proteins XPA and XPC/HR23B to the 5R-thymine glycol lesion and structure of the cis-(5R,6S) thymine glycol epimer in the 5′-GTgG-3′ sequence: destabilization of two base pairs at the lesion site

The 5R thymine glycol (5R-Tg) DNA lesion exists as a mixture of cis-(5R,6S) and trans-(5R,6R) epimers; these modulate base excision repair. We examine the 7:3 cis-(5R,6S):trans-(5R,6R) mixture of epimers paired opposite adenine in the 5′-GTgG-3′ sequence with regard to nucleotide excision repair. Human XPA recognizes the lesion comparably to the C8-dG acetylaminoflourene (AAF) adduct, whereas XPC/HR23B recognition of Tg is superior. 5R-Tg is processed by the Escherichia coli UvrA and UvrABC proteins less efficiently than the C8-dG AAF adduct. For the cis-(5R, 6S) epimer Tg and A are inserted into the helix, remaining in the Watson–Crick alignment. The Tg N3H imine and A N6 amine protons undergo increased solvent exchange. Stacking between Tg and the 3′-neighbor G•C base pair is disrupted. The solvent accessible surface and T2 relaxation of Tg increases. Molecular dynamics calculations predict that the axial conformation of the Tg CH3 group is favored; propeller twisting of the Tg•A pair and hydrogen bonding between Tg OH6 and the N7 atom of the 3′-neighbor guanine alleviate steric clash with the 5′-neighbor base pair. Tg also destabilizes the 5′-neighbor G•C base pair. This may facilitate flipping both base pairs from the helix, enabling XPC/HR23B recognition prior to recruitment of XPA

Oxidative DNA Damage and Repair: Mechanisms, Mutations, and Relation to Diseases

Oxidative DNA damage (ODD) by reactive oxygen species (ROS) or reactive nitrogen species (RNS) is an inevitable tradeoff for using oxidation processes by living cells as a source of energy [...

Selective Radiation-Induced Generation of 2-Deoxyribonolactone Lesions in DNA Mediated by Aromatic Iodonium Derivatives

2-Deoxyribonolactone lesions were identified as major products of radiation damage to DNA mediated by o,o\u27-di-phenyleneiodonium cations in a hydroxyl radical-scavenging environment. The highest selectivity toward deoxyribonolac-tone formation (up to 86% of all sugar-phosphate damages) and the overall reaction efficiency (up to 40% of all radiation-generated intermediates converted into products) was displayed by derivatives with positively charged (2-aminoethyl-thio)acetylamino and (2-aminoethylamino)acetylamino side chains. The reaction can be useful for random single-step incorporation of deoxyribonolactone lesions into single- and double-stranded oligonucleotides and highly polymerized DNA directly in commonly used buffers (PBS, phosphate, Tris-HCl, etc.) at room temperature. In combination with HPLC separation, this technique can serve as a source of short (\u3c6 mer) sequences containing deoxyribonolactone lesions at known positions

Association with Polyamines and Polypeptides Increases the Relative Yield of 2-Deoxyribonolactone Lesions in Radiation-Damaged DNA

The production of 2-deoxyribonolactones (C1\u27-oxidation product), C4\u27-oxidized abasic sites and C5\u27-carbonyl terminated strand scission products was investigated in complexes of double-stranded DNA with protamine, poly-L-lysine and spermine exposed to X-ray radiation. The lesions were quantified by high-performance liquid chromatography through the release of the corresponding low-molecular-weight products 5-methylenefuran-2(5H)-one, N-(2\u27-hydroxy-ethyl)-5-methylene-D3-pyrrolin-2-one and furfural, respectively. All binders were found to increase the relative yield of C1\u27 oxidation up to 40% of the total 2-deoxyribose damage through the indirect effect versus approximately 18% typically found in homogeneous solutions by the same technique. On the contrary, the yield of C5\u27-oxidation was found to be suppressed almost completely, while in homogeneous solutions it constituted approximately 14% of the total. The observed change in end product distribution is attributed to free valence transfer to and from the complexing agent, although the mechanisms associated with this process remain unclear

2-Deoxyribonolactone Lesions in X-Ray-Irradiated DNA: Quantitative Determination by Catalytic 5-Methylene-2-Furanone Release

(Chemical Equation Presented) Torn genes: DNA tetramers containing the 2-deoxyribonolactone (dL) lesion have been isolated by HPLC from d(CGCG) and d(pCGCG) films irradiated with X-rays. Upon treatment with spermine as a catalyst, the dL-containing tetramers decompose to 5-methylene-2-furanone (5-MF; see scheme), a characteristic product of dL decomposition. Hence, 5-MP can be used to quantify dL lesions in DNA

Diffusion Approach to Long Distance Charge Migration in DNA: Time-Dependent and Steady-State Analytical Solutions for the Product Yields

In this study we report analytical solutions for both time-dependent and steady-state problems of unbiased charge transfer through a regular DNA sequence via a hopping mechanism. The phenomenon is treated as a diffusion of charge in a one-dimensional array of equally spaced and energetically equivalent temporary trapping sites. The solutions take into account the rates of charge hopping (k), side reactions (kr), and charge transfer to a terminal charge acceptor (kt). A detailed analysis of the time-dependent problem is performed for the diffusion-controlled regime, i.e., under the assumption that kt ≫ k, which is also equivalent to the fast relaxation limit of charge trapping. The analysis shows that the kinetics of charge hopping through DNA is always multiexponential, but under certain circumstances it can be asymptotically approximated by a single-exponential term. In that case, the efficiency of charge transfer can be characterized by a single rate constant kCT = 1.23kN-2 + kr, where N is the DNA length expressed in terms of the number of equidistant trapping sites and kr is the rate of competing chemical processes. The absolute yield of charge transfer under steady-state conditions in general is obtained as Y∞ = ω[α sinh(αN) + ω cosh(αN)]-1, where α = (2kr/k)1/2 and ω = 2kt/k. For the diffusion-controlled regime and small N, in particular, it turns into the known algebraic dependence Y∞ = [1 + (kr/k)N2]-1. At large N the solution is asymptotically exponential with the parameter α mimicking the tunneling parameter β in agreement with earlier predictions. Similar equations and distance dependencies have also been obtained for the damage ratios at the intermediate and terminal trapping sites in DNA. The nonlinear least-squares fit of one of these equations to experimental yields of guanine oxidation available from the literature returns kinetic parameters that are in reasonable agreement with those obtained by Bixon et al. [Proc. Natl. Acad. Sci. U.S.A. 1999, 96, 11713-11716] by numerical simulations, suggesting that these two approaches are physically equivalent

Protection of DNA Against Direct Radiation Damage by Complex Formation With Positively Charged Polypeptides

Radioprotection of DNA from direct-type radiation damage by histones has been studied in model systems using complexes of positively charged polypeptides (PCPs) with DNA. PCPs bind to DNA via ionic interactions mimicking the mode of DNA-histone binding. Direct radiation damage to DNA in films of DNA-PCP complexes was quantified as unaltered base release, which correlates closely with DNA strand breaks. All types of PCPs tested protected DNA from radiation, with the maximum radioprotection being approximately 2.5-fold compared with non-complexed DNA. Conformational changes of the DNA induced by PCPs or repair of free radical damage on the DNA sugar moiety by PCPs are considered the most feasible mechanisms of radioprotection of DNA. The degree of radioprotection of DNA by polylysine (PL) increased dramatically on going from pure DNA to a molar ratio of PL monomer:DNA nucleotide ∼1:2, while a further increase in the PL:DNA ratio did not offer more radioprotection. This concentration dependence is in agreement with the model of PCP binding to DNA that assumes preferential binding of positively charged side groups to DNA phosphates in the minor groove, so that the maximum occupancy of all minor-groove PCP binding sites is at a molar ratio of PCP:DNA = 1:2