Direct Ionization of DNA

Since there is such an imprecise division between direct and indirect effects in the literature, some experimental results are presented to clarify this classification. Basically, one cannot detect HO radicals at low DNA hydrations (ca. 10 water molecules per nucleotide) [12]. This means that in the first step of ionization, the hole produced in the DNA hydration shell transfers to the DNA. It is impossible to distinguish the products from the hole or electron initially formed in the water from the direct-effect damage products. For this discussion, direct-type damage will be considered to arise from direct ionization of DNA or from the transfer of electrons and holes from the DNA solvation shell to the DNA itself. 

Brief Review of Studies of Electron and Hole Formation and Transfer After Direct Ionization of DNA... 

Scheme 5.18 Product formation by direct ionization of DNA at exposure to high-energy radiation. B, base dR, deoxyribose [87].

Ionization of DNA s solvation shell produces water radical cations (H20 ) and fast electrons. The fate of the hole is dictated by two competing reactions hole transfer to DNA and formation of HO via proton transfer. If the ionized water is in direct contact with the DNA (F < 10), hole transfer dominates. If the ionized water is in the next layer out (9 < r < 22), HO formation dominates [67,89,90]. The thermalized excess electrons attach preferentially to bases, regardless of their origin. Thus the yield of one-electron reduced bases per DNA mass increases in lockstep with increasing F, up to an F of 20-25. This means that when F exceeds 9, there will be an imbalance between holes and electrons trapped on DNA, the balance of the holes being trapped as HO . At F = 17, an example where the water and DNA masses are about equal, the solvation shell doubles the number of electron adducts, increasing the DNA-centered holes by a bit over 50% [91-93]. 

A discussion of the distribution of stable end products produced by direct ionization in DNA is presented. Single- and double-strand break yields for low LET radiation are discussed. Then work on end products derived from the DNA bases is reviewed. Recent... 

DNA suffers radiation damage by both a direct mechanism and an indirect mechanism. The direct damage results from radiation-induced ionization of DNA itself. The indirect damage results from attack on DNA by other free radicals. Since water is the predominant molecule in cells, the major source of indirect radiation DNA damage is from water radiolysis. 

Bernhard and co-workers have performed a series of experiments to determine the mechanisms of DNA strand breakage by direct ionization of plasmid DNA. A big surprise in this work was the discovery that the total yield of single strand breaks exceeds the yield of trapped sugar radicals. Even at very low hydration levels (2.5 waters per nucleotide residue) nearly 2/3 of the strand breaks are derived from precursors other than deoxyribose radicals [74], The authors conclude that a majority of the strand breaks observed do not result from dissociative electron capture, homolytic bond cleavage from excited states, or from hydroxyl radical attack. Rather, the authors conclude that doubly oxidized deoxyribose is responsible for the high yield of strand breaks. 

As described earlier, formation of substantially increased amounts of DNA-backbone-sugar and -phosphate radicals are observed in argon and oxygen ion-beam irradiated hydrated DNA samples relative to those found with low LET / irradiation.The usual mechanism suggested for formation of sugar radicals, i.e. hole deprotonation from direct ionization of the cannot explain the... 

Purkayastha S, Milligan JR, Bernhard WA. (2006) An investigation into the mechanisms of DNA strand breakage by direct ionization of variably hydrated plasmid DNA. J Phys Chem B 110 26286-26291. 

The double-stranded structure of DNA not only affects the chemistry of OH-induced damage but also may modulate oxidation reactions that take place upon initial direct ionization of a given nucleotide. Indeed, the positive holes created in the latter process have... 

Douki T., Ravanat J.-L., Pouget J.-P, Testart I., Cadet J., Minor contribution of direct ionization to DNA base damage induced by heavy ions, Int. J. Radiat. Biol., 2006,82, 119-127. 

In contrast to the direct induction of DNA damage by UV-C and UV-B light, UV-A produces damage indirectly through highly reactive chemical intermediates. Similar to ionizing radiation, UV-A generates... 

The ability to detect small genetic changes becomes more difficult as mass increases. There is further an upper mass range where analysis is impractical. For low-resolution instruments this limit is around a 100 mer. Thus the mass has to be minimized or a high-resolution instrument employed. Alternatively, the smaller the piece of DNA analyzed, the more it chemically resembles a primer or nucleotide monomer thus separation of the two during cleanup is difficult to do. If the primers and nucleotides are not removed, they can provide a massive background on MS analysis or inhibit ionization of the PCR product by preferential ionization. Thus for practical reasons it is extremely difficult to employ a PCR product below a 40 to 50mer for direct ESI MS or ESI MS-MS analysis. 

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