Photolyases

Flavins — Riboflavin is first of all essential as a vitamin for humans and animals. FAD and FMN are coenzymes for more than 150 enzymes. Most of them catalyze redox processes involving transfers of one or two electrons. In addition to these well known and documented functions, FAD is a co-factor of photolyases, enzymes that repair UV-induced lesions of DNA, acting as photoreactivating enzymes that use the blue light as an energy source to initiate the reaction. The active form of FAD in photolyases is their two-electron reduced form, and it is essential for binding to DNA and for catalysis. Photolyases contain a second co-factor, either 8-hydroxy-7,8-didemethyl-5-deazariboflavin or methenyltetrahydrofolate. ... 

Folate and FAD are also components of cryptochromes, proteins widespread in living organisms. Cryptochromes are considered photolyase sequence homologues with no DNA repair activities but with blue light-activated factors. Cryptochromes regulate growth and development in plants and seem to be responsible for the synchronization of circadian rhythms in animals and human. ... 

Sancar, A., Structure and function of DNA photolyase and cryptochrome blue-light... 

Electron donation to nucleobases is a fundamental process exploited by nature to achieve the efficient repair of UV induced lesions in DNA [27, 28]. Nature developed to this end two enzymes, CPD photolyases and (6-4) photolyases, which both inject electrons into the UV damaged DNA bases [29, 30]. Both enzymes are, in many species, including plants, essential for the repair of the UV-light induced DNA lesions depicted in Scheme 1 [31]. 

Although the reduction potentials of DNA bases and UV induced DNA lesions inside a DNA double strand or inside the active site of a DNA photolyase, together with the reduction potential of the photoexcited FADH- in the photolyases, are not known, currently available redox potentials indicate that the single electron reduction of a nucleobase or a UV induced dimer lesion by a reduced and deprotonated flavin coenzyme is a weakly exothermic process. The reduced and deprotonated FADH- in its photoexcited state is... 

Scheme 2 Mechanism of repair of cyclobutane pyrimidine dimers (CPD) by a CPD photolyase. 8-HDF 8-hydroxy-5-deazaflavin, ET electron transfer. FADH reduced and de-protonated flavin-coenzyme...

In order to investigate the single electron donation process from a reduced flavin to a pyrimidine dimer or oxetane lesion, the photolyase model compounds 1-4 depicted in Scheme 4 were prepared [41, 42]. The first model compounds 1 and 2 contain a cyclobutane uracil (1) or thymine (2) dimer covalently connected to a flavin, which is the active electron donating subunit in photolyases. These model systems were dissolved in various solvents... 

These experiments proved that a light-excited, reduced flavin is indeed able to photoreduce cyclobutane pyrimidine dimers and that these dimers undergo a spontaneous cycloreversion. The quantum yield of about 0=5% clarified that the overall dimer splitting process is highly efficient, even in these simple model systems ((]) photolyase 70%). 

Flavin-cyclobutane pyrimidine dimer and flavin-oxetane model compounds like 1-3 showed for the first time that a reduced and deprotonated flavin is a strong photo-reductant even outside a protein environment, able to transfer an extra electron to cyclobutane pyrimidine dimers and oxetanes. There then spontaneously perform either a [2n+2n cycloreversion or a retro-Paternd-Buchi reaction. In this sense, the model compounds mimic the electron transfer driven DNA repair process of CPD- and (6-4)-photolyases. 

A computational study was concerned with the effect of solvation on the radical ion involved in CDP photolyase enzyme-catalysed reversion of thymine and uracil cyclobutane dimers stimulated by visible light <06T6490>. 

An in-depth study of DNA repair systems (Aravind et al., 1999a) has concluded that few, if any, repair proteins occur with identical collinear domain arrangements in all three kingdoms of life. Approximately 10 enzyme families of adenosine triphosphatases (ATPases), photolyases, helicases, and nucleases were identified that are all likely to have been present in the cenancestor. These enzymatic domains are accompanied in DNA repair proteins by numerous regulatory domains. This indicates that the domain architectures of these proteins are labile, with incremental addition and/or subtraction of domains to conserved cores to be a common phenomenon except in the most closely related species. 

Carell has recently presented the study of a flavin amino acid chimera to model riboflavin in DNA photolyases [68]. This amino acid LI (Fig. 20) was synthesized in an enantiopure fashion by building the alloxazine ring onto the epsilon amine of lysine. This coenzyme chimera was applied to the problem of repairing DNA damage caused by UV irradiation. LI was incorporated into an 21-residue peptide, P-1, possessing the sequence of the DNA-binding domain of the helix-loop-helix transcription factor MyoD. 

Gaillard, H., Fitzgerald, D.J., Smith, C.L., Peterson, C.L., Richmond, T.J., and Thoma, F. (2003) Chromatin remodeling activities act on UV-damaged nucleosomes and modulate DNA damage accessibility to photolyase. J. Biol. Chem. 278, 17655-17663. 

Thymine dimers can be removed by photoreactivation (2). A specific photolyase binds at the defect and, when illuminated, cleaves the dimer to yield two single bases again. 

Sancar GB, Jorns MS, Payne G, Fluke DJ, Rupert CS, Sancar A 1987a Action mechanism of Escherichia coli DNA photolyase. III. Photolysis of the enzyme-substrate complex and the absolute action spectrum. J Biol Chem 262 492-498 Sancar GB, Smith FW, Reid R, Payne G, Levy M, Sancar A 1987b Action mechanism of Escherichia coli DNA photolyase. I. Formation of the enzyme-substrate complex. J Biol Chem 262 478-485... 

Van Gelder That is the photolyase model. In photolyase, the damaged DNA is bound in the dark, and then dissociates following the light-mediated enzymatic repair of the thymidine dimer. Photolyases are believed to be the molecular ancestors of cryptochromes. 

Herzog ED, Takahashi JS, Block GD 1998 Clock controls circadian period in isolated suprachiasmatic nucleus neurons. Nat Neurosci 1 708—71 KobayashiK, Kanno S, Smit B, van der Horst GTJ, Takao M, Yasui A 1998 Characterization of photolyase/blue-hght receptor homologs in mouse and human cells. Nucleic Acids Res 26 5086-5092... 

Todo T, Ryo H, Yamamoto K et al 1996 Similarity among the Drosophila (6-4)photolyase, a human photolyase homolog, and the DNA photolyase-blue-Kght photoreceptor family. Science 272 109-112... 

Kay We have to be very careful about cryptochrome. Cashmore has made the observation that he thinks the cryptochromes are more similar to their own photolyases than to each other, and has used this observation to suggest that cryptochromes have each arisen independently. It is not surprising that cryptochromes could have arisen independently and still be involved in clocks. One of the most stressful stimuli a cell can be exposed to is light. You can imagine one of the first clocks being built around fight dependency of DNA repair. It is wrong to think of cryptochromes as a conserved element of clocks because they could have arisen independently. 

FIGURE 8.2 Formation of a thymine-thymine dimer by UV-B radiation, and repair by UV-A or blue light-activated photolyase. 

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