Cytosine deprotonated

Summary. The studies of cytosine monohydrate doped with 2-thiocytosine find evidence for hole transfer from cytosine to thiocytosine. If transfer is considered to occur along only the a-stacked bases, 31 relatively large transfer distances are calculated. In more recent work, however, the authors suggests that the transfer is more likely to be 3-dimensional in character and, thus, the overall transfer distance is only a few bases at 15 K.33 The authors suggest the trapping of the hole on cytosine is accompanied by deprotonation which stabilizes the hole on this site and prevents further transfer and that hole transfer is competitive process with cytosine deprotonation. A hypothesis is put forward that the ill understood 3aH radical may originate with small amounts of 5-methylcytosine in the systems studied. 

Figure 3.12 depicts TOP SIMS spectra obtained from ODN and PNA immobilized on silanized silicon wafers. The spectra clearly demonstrate that the masses corresponding to POi and PO3 provide the best correlation of the presence of ODN, enabling their use for precise distinction between ODN and PNA. The CFJ and C2O2FJ peaks seen in the PNA spectra represent trifluoroacetic acid, which was part of the PNA solution. Deprotonated (Cyt-H) and (Thy-H) signals of the bases cytosine and thymine are observed for both immobilized PNA and ODN sequences and can be used to detect the presence of these bases. 

Deoxycytidine (dCyd) (14 in Scheme 2) is also an excellent target for one-electron oxidation reactions mediated by triplet excited menadione. On the basis of extensive identification of dCyd photooxidation products, it was concluded that this nucleoside decomposes by competitive hydration and deprotonation reactions of cytosine radical cations with yields of 52% and 40%, respectively [53]. It was also found, on the basis of 180 labeling experiments, that hydration of cytosine radical cations (15) predominantly occurs... 

Deprotonation provides the necessary electron push to kick out the electron pair joining C(6) with the nitrobenzene oxygen. If, however, N(l) is alkylated (as with the nucleosides and nucleotides), OH catalysis is much less efficient since it now proceeds by deprotonation from N(3) (with the uracils) or from the amino group at C(4) (with the cytosines). In these cases the area of deprotonation is separated from the reaction site by a (hydroxy)methylene group which means that the increase in electron density that results from deprotonation at N(3) is transferable to the reaction site only through the carbon skeleton (inductive effect), which is of course inefficient as compared to the electron-pair donation from N(l) (mesomeric effect) [26]. Reaction 15 is a 1 1 model for the catalytic effect of OH on the heterolysis of peroxyl radicals from pyrimidine-6-yl radicals (see Sect. 2.4). 

Another example relates to OH -aided one-electron oxidation of cytosine. With cytosine, the OH reaction proceeds by addition to C(5), a process that has a selectivity of 90% [24]. The 5-hydroxy-6-yl radical is an excellent reductant, and the same is true for the ionized 6-yl radical formed by deprotonation from N(l). This radical anion now contaim sufficient electron density to eliminate the OH group at C(5) as OH". The result is the cytosine-l-yl radical which is oxidizing, probably due to appreciable spin density at the hetero atoms N(l) and... 

These ideas have been illustrated in a recent study of the co-crystalline complex of 1-meCyt 5-FUra [19]. Using model calculations, it was shown how the hydrogen-bonding network of the crystal is able to sustain a proton shuttle which leads to the selective formation of certain radicals. Calculations predict that the site of reduction would be the cytosine base, yielding the N3 protonated cytosine anion, Cyt(N3-I-H), while the uracil base would be the site of oxidation, yielding the N1 deprotonated uracil cation, Ura(Nl—H) ... 

Oxidation of cytosine produces a radical with sites of unpaired spin density at Nl, N3, and C5. The cytosine cation has a pKn<4 and in solution deprotonates at NH2 [22]. In the solid state, Sagstuen et al. [24] assigned the primary oxidation radical observed in cytosine monohydrate as the Nl deprotonated cation, Cyt(Nl—H) . It is known from the ENDOR... 

Some time ago, an allyl-like radical was observed in irradiated crystals of 5 -dCMP [26]. This radical was thought to be a sugar radical, although no likely scheme was proposed for its formation. It now appears that this radical is formed on 5-methyl cytosine impurities in these crystals [27]. This radical forms by deprotonation at the methyl group of the cytosine cation, 5meCyt(Me—H) , and may have important consequences in the radiation chemistry of DNA since the ionization potential of 5-methyl cytosine is lower than that of either cytosine or thymine. 

The presence of such a mechanism was prohed hy varying the sweep rate as illustrated in Fig. 4. Caution must he exercised in drawing conclusions from such a study because, with hahde ions (and with some other unidentate species), bridging by the halide between the copper ion and the electrode surface may accelerate the rate of electron transfer and lead to erroneous conclusions. This type of mechanism has also been proposed by Palaniandavar and coworkers [121] for the Cu(II/I) complex with deprotonated salicylideneglycine in the presence of cytosine or cytidine in which the latter species tends to be coordinated only to the oxidized complex. 

In double-stranded DNA, electron abstraction from the guanine radical cation can be associated with an extremely fast shift of the N1 proton to its Watson-Crick partner cytosine (Scheme 2a) [9]. The equilibrium constant for the protonation of C (pfCa=4.3) with the concomitant deprotonation of G estimated from the pK values of the free nucleosides, is about 2.5 [49]. Within these constraints, the guanine radical should retain some radical cation character [82] and the complete deprotonation of G would require a base pair opening event occurring on a millisecond timescale [74]. An alternative mechanism of G deprotonation is the release of the N2 proton (Scheme 2b). This mechanism was experimentally established for 1-methyl-guanosine conductometric results showed that in neutral solutions, the radical cation of this nucleoside rapidly deprotonates with the formation of the neutral radical [48]. Although the exact mechanism of the G deprotonation in double-stranded DNA requires further clarification, electron abstraction... 

Some other interesting observations regarding free radicals in these systems are noteworthy. In many instances, multiple conformations of radicals are found at lower but not higher temperatures. This indicates that the radicals exist in shallow energy wells at low temperature this phenomenon was observed very early, in the 4 K ENDOR investigation of radical formation in amino acids.23 Unlike the process in DNA. In which it is well understood that the thymine anion radical protonates at C6 to form T(C6)H-, in the crystalline state there is a not clear link between pyrimidine electron adducts and H-addition radicals. We finally note that a deuterium isotope effect of protonation/deprotonation processes was found in cytosine.HCl and 2 -deoxycytidine.HCl, as evidenced by a lower propensity for these processes to occur in partially deuterated systems than in predated ones. 

Another recent work looks at possible hole transfer in crystals of cytosine HC1 doped with 5-methylcytosine HCl.34 By controlling the amount of dopant (5-methylcytosine) a measure of hole transfer through the crystal is found. Irradiation forms both the cytosine and the 5-methylcytosine cation radicals, the later of which decays by deprotonation to the 3aH radical. Far more of this radical is found at elevated temperatures than at 77 K, or on annealing crystals irradiated at low temperatures to room temperature. The authors suggest that hole transfer and deprotonation to form the 3aH radical is an activated process which is fast at room temperature and slow at lower temperatures where little evidence for transfer is found. 

A comparison of aliphatic amides, cyclic amides, cyclic imides and 2,4-dioxopyrimidines (uracils) in their deprotonated and diplatinated form (Scheme 4) reveals an increasing steric shielding of the V-bonded Pt ion (Ptx). With respect to formation of stacked and partially oxidized dinuclear species, it is evident that application of the binding principles seen in the blues of cyclic amides to the uracils and imides allows for tetranuclear species only. On the other hand, the presence of an additional O-donor in the imides and uracils (and likewise the cytosines, vide infra) provides an... 

On the other hand, electrophilic metal binding at C(5) with consequent deprotonation of this site was observed [12], Mono- (at C(5)) [18-20] and di-deprotonated species (C(5),N(4)) [21] have been reported. The shorthand used to represent deprotonation at C(5) for uracils and cytosines bears the negative charge before the shown AT-bonded H-atoms. In the Table, the shorthand for these anions is reported in italics. 

The uracilate and thyminate anions simultaneously bind the metals through N(3) and 0(4), whereas the 1-methylcytosinate anions do so through N(3) and the deprotonated amino group N(4) [14]. The neutral cytosine was found also to act as bridging ligand through N(3) and 0(2) [33]. 

Since an oxidation product on cytosine in DNA could not deprotonated at Nl, it may be more relevant to look at oxidation in a nucleotide. In 5 -dCMP (with N3-H in the native molecule) oxidation produces the N3 deprotonated cation with p(Nl) = 0.30 and p(C5) = 0.60 [43],... 

Wetmore et al. also examined the oxidation product in cytosine [79], They computed spin densities p(Nl) = 0.29 and p(C5) = 0.49 for the N1 deproto-nated cation observed in cytosine monohydrate. These results are very close to the experimental results presented in Section 18.4.1.1 p(Nl) = 0.30 and p(C5) = 0.57. However, since their calculated C5-H isotropic hyperfine coupling (—31.5 MHz) is significantly different from the experimental value (—41.4 MHz), and their calculation predicts only a small N4 spin density, they reject the N1 deprotonated cation model. To see why this is not correct, one can invoke the litany of observations presented above from a radiation chemistry perspective. 

First of all the high spin density on C5 is indicative of an oxidation product. In order to be stably trapped, cations have to deprotonate. In cytosine monohydrate, this deprotonation can most easily occur at N1 or N4. Deprotonation at the amino group would give a radical species that would not fit the EPR/ENDOR data. Therefore the N1 deprotonated cation is the best model to represent the experimental data, and actually the best model from the calculations that Wetmore et al. performed [79], The disagreement Wetmore et al. report is with the C5-H isotropic hyperfine coupling. This is actually to be expected since the authors have not included the important effects of the hydrogen bonded network present in the single crystal in their calculations. 

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