Irradiation nucleic acids

WHO (1993) Guidelines for Drinking Water Quality, 2nd Ed., Vol. 1. Recommendations, Geneva Wilmer, J.W.G.M. Natarajan, A.T. (1981) Induction of sister-chromatid exchanges and chromosome aberrations by y-irradiated nucleic acid constituents in CHO cells. Mutat. Res., 88,... 

In recent years there has been much interest in the chemistry and biochemistry of irradiated nucleic acids and their derivatives (B-76MI40901). ESR spectroscopy is concerned with the structure of radical forms produced by high energy radiation and has been of value in understanding the complex reactions which occur in the nucleic acids and their simpler derivatives. A discussion of these last reactions is beyond the scope of this chapter and the reader is referred to the bibliography in Table 2. 

J.F. Ward, Chemical consequences of irradiating nucleic acids, J. Agric. Food Chem., 26... 

Very important information concerning the photochemistry of the nucleic acids was furnished by the report of Beukers and Berends that irradiation of frozen solutions of thymine produces a stable photoproduct corresponding to dimerization of the thymine/58 This photoproduct has been subsequently identified as a cis-syn-cis dimer ... 

It is well known that pyrimidine bases convert to photodimers upon irradiation to UV light near the X max( > 270 nm). This photochemical reaction has a lethal effect in biological systems due to the photochemical transformation of pyrimidine bases of nucleic acids. However the photodimerization is a reversible reaction and the photodimers split to afford the original monomers very efficiently upon irradiation at a shorter wavelengths as shown in Scheme 1(1). 

Thymine and other bases in nucleic acids undergo dimerisation when irradiated with ultraviolet light, and this can result in DNA mutation or even cell death. 

Orotic acid readily forms dimers even when irradiated in liquid medium [582, 583]. 5-Bromouracil (5-BrU) in DNA is dehalogenated, rather than forming cyclobutane-type dimers. Such DNA derivatives are more sensitive to ultraviolet irradiation than normal DNAs [584-594], Irradiation of 5-bromo-uracil and derivatives in aqueous medium produces 5,5 -diuracil [590, 591]. However, derivatives such as 3-sbutyl-5-bromo-6-methyluracil have been reported to yield cyclobutane dimers either by irradiation of frozen aqueous solutions, or by catalysis with free radical initiators, such as aluminium chloride, ferric chloride, peroxides or azonitriles [595]. 5-Hydroxymethyluracil is reported to dimerize very slowly in frozen water at 2537 A [596]. The fundamental research in the photochemistry of the nucleic acids, the monomeric bases, and their analogues has stimulated new experiments in certain micro-organisms and approaches in such diverse fields as template coding and genetic recombination [597-616]. 

The previous section outlined the typical e loss and e gain products observed in the nucleic acid bases in the solid state. These studies can be applied to the study of the radiation chemistry of DNA. The relevance of the study of model systems is shown by considering the following remarkable observations. Years ago, Ehrenberg et al. showed the EPR spectra of the 5,6-dihydrothymine-5-yl radical observed in thymine, thymidine, and DNA. The spectra are nearly identical [46]. The reduction product observed in cytosine monohydrate is the N3 protonated anion. In solution, this reduction product gives rise to a 1.4-mT EPR doublet. The same feature is present in irradiated DNA at 77 K. Likewise, the result of e loss in guanine bases is characterized by a broad EPR singlet. The same feature is also evident in the EPR spectrum of DNA irradiated and observed at 77 K. 

Another method is to produce EPR basis spectra by irradiating various nucleic acid bases. The EPR spectrum of DNA is simulated by taking various combinations of the... 

This review has looked at the direct effects of ionizing radiation on nucleic acids. The first step was to review detailed EPR/ENDOR experiments on irradiated model compounds at low temperatures in order to study the primary radiation-induced defects. Next, it was shown how these EPR spectra are used to simulate the EPR spectra of the DNA polymer. 

Although a signal was detected also in irradiated pyrimidine solutions under similar conditions, it was attributed to dissociation products of the pyrimidine molecule and not to photoionization. If the purine solutions contained ethanol, then the signal of the CH3CHOH radical could also be detected this did not happen in ethanolic pyrimidine solutions. The possible importance of photoionization as the primary process in the photochemistry of nucleic acid derivatives has been supported by Kearns, from a study of photoionization in anthracene single crystals.115 He concluded that anthracene photoionization occurred via the interaction of two singlet excitons. 

Also interesting is the work hy Franchi and Gallon (Franchi et al., 2003 Franchi and Gallon, 2004), who showed that once nucleic acids are adsorbed on clay, they are much more resistant to irradiation, temperature shock, and other extreme conditions. 

An unusual photochemical reaction of 2-pyridones, 2-aminopyridinium salts and pyran-2-ones is photodimerization to give the so-called butterfly dimers. These transformations are outlined in equations (13) and (14). Photodimerization by [2+2] cyclization is also a common and important reaction with these compounds. It has been the subject of particular study in pyrimidines, especially thymine, as irradiation of nucleic acids at ca. 260 nm effects photodimerization (e.g. equation 15) this in turn changes the regular hydrogen bonding pattern between bases on two chains and hence part of the double helix structure is disrupted. The dimerization is reversed if the DNA binds to an enzyme and this enzyme-DNA complex is irradiated at 300-500 nm. Many other examples of [2+2] photodimerization are known and it has recently been shown that 1,4-dithiin behaves similarly (equation 16) (82TL2651). 

All these bases absorb around 260 nm. Thymine and cytosine are most sensitive to irradiation. Two most important types of photochemical reactions that have been observed for these pyrimidine bases are photohydration and photodimerization. In vivo systems, interactions between protons and nucleic acids can also be initiated by radiations of wavelength dlorter than 300 nm. 

Photochemically reactive molecules have often been used as labels for specific sites in proteins and nucleic acids. Psoralen derivatives serve as relatively nonspecific photochemically activated crosslinking agents for DNA and double-stranded RNA.195 Aryl azides are converted by light to aryl nitrenes, which react in a variety of ways including insertion into C-H bonds (Eq. 23-27).200 201 In some cases UV irradiation can be used to join natural substrates to enzymes or hormones to receptors. For example, progesterone, testosterone, and other steroids have been used for direct photoaffinity labeling of their receptors.202 Synthetic benzophenones have also been used widely as photoactivated probes.203... 

Nucleic acids can be visualized on the slab gel after separation by soaking in a solution of ethidium bromide, a dye that displays enhanced fluorescence when intercalated between stacked nucleic acid bases. Ethidium bromide may be added directly to the agarose solution before gel formation. This method allows monitoring of nucleic acids during electrophoresis. Irradiation of ethidium bromide-treated gels by UV light results in orange-red bands where nucleic acids are present. 

Chemically modified DNAs can also be used as hybridization probes, provided that the modification does not interfere with the formation of hybrid DNA molecules. A psoralen biotin label has also been developed. Psoralen is a photoactivable agent that can intercalates into single- or double-stranded nucleic acids. On irradiation at 365 nm, it will covalently bind to the probes. This labeling reaction is simple and straightforward. However, the reagents for labeling and detection are only available in a kit format. 

That direct irradiation with ultraviolet light is damaging to single-cell organisms is well known. It also is known that the nucleic acids, DN A and RN A, are the important targets of photochemical damage, and this knowledge has stimulated... 

An interesting and significant outcome is the finding that the pyrimidine bases of nucleic acids (uracil, thymine, and cytosine) are photoreactive and undergo [2 + 2] cycloadditions on irradiation with ultraviolet light. Thymine, for example, gives a dimer of structure 7 ... 

---