Enzymes photolyases

A number of studies are concerned with the free-radical reactions of typical nucleobase lesions. For example, the cyclobutane-type Thy dimer can be split by one-electron reduction [Heelis et al. 1992 reactions (307) and (308)], a process that is relevant to the repair of this typical UV-damage by the photoreactivating enzyme (photolyase, for a review see Carrell et al. 2001, for the energetics of the complex reaction sequence, see Popovic et al. 2002). At 77 K, the dimer radical anion is sufficiently long-lived to be detectable by EPR (Pezeshk et al. 1996). 

The light-induced repair of cylobutane-type dimers by the enzyme photolyase is of major biological importance. This proceeds by ET from a flavin, and using a model system it has been shown that the electron is likely to be funneled through the DNA base stack (SchwOgler et al. 2000). 

In direct repair the chemical modification that constitutes the lesions is reversed without removing and replacing nucleotides. There are four known direct repair enzymes photolyase, spore photoproduct lyase, methylguanine DNA methyltransferase, and AUcB family oxidative demethylases (1). 

FIGURE 2.14. Formation of dimers by UV radiation and their photoreactivation by the enzyme photolyase. 

Cryptochrome has a high level of sequence identity to the blue-hght activated enzyme, photolyase. 

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. ... 

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]. 

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. 

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... 

There has been considerable interest in the mechanism of action of DNA photolyase enzymes, which repair pyrimidine dimers in damaged DNA via action of visible light. Several model reactions have given credence to a possible SET pathway. For example, Falvey and coworker reported that the photochemical reaction of dimer 35 with FADH2 results in a retro [2 + 2] cycloaddition according to Scheme 2269. 

Several amphibian species were examined for photolyase activity. This enzyme is responsible for the repair of DNA damage caused by UV-B radiation. A more than... 

Thymidine cyclobutane dimers are important photoproducts formed by short-wave UV irradiation (2 = 290-320 nm) of DNA, by [2 + 2] cycloaddition between two adjacent thymine nucleobases in the same oligonucleotide strand (Scheme 4.5.1) [1]. They lead to profound biological effects in vivo, including mutation, cancer, and cell death [2] (Box 21). In a wide range of organisms the repair of these lesions in DNA is accomplished by enzymes (the photolyases), which regenerate undamaged thymidines by means of a photoinduced electron-transfer process [3]. 

Photoreaction can remove thymidine-thymidine photodimers. Visible light activates the enzyme DNA photolyase. This enzyme absorbs visible light and transfers that energy to the photodimer, reversing the dimerization directly. 

Several natural photoenzymes with activity controlled by light have been reported to date. Among them are DNA and (6-4) photoproduct photolyases, which are highly efficient light-driven DNA-repair enzymes [14, 15], protochlorophyllide reductase, which is an important enzyme in the chlorophyll biosynthesis pathway [16, 17], nitrile hydratase, which hydirates aliphatic and aromatic nitriles to the... 

Sanders DB, Wiest O. A model for the enzyme-substrate complex of DNA photolyase and photodamaged DNA. J Am Chem Soc 1999 121 5127-34. 

Belanich et al.63 reported the removal of endotoxins from protein mixtures. Endotoxins in a bacterial extract containing a protein photolyase was passed through a stack of 10 disk membranes (Q-type, Sartorius). LAL assay was used to monitor the endotoxin levels after each pass. There was over 5 log reduction in endotoxin content after three passes through the membranes. The protein content was reduced during this process, however, the enzyme s specific activity was increased 35-fold. This study also determined that the binding capacity of the membrane was greater than 2.25 million EU/cm2 of membrane area. 

Figure 23.1. Schematic illustration of direct reversal of a pyrimidine dimer by the enzyme DNA photolyase. The enzyme binds to the pyrimidine dimer present in DNA. The square and triangle represent the two noncovalently bound chromophores that are present in all photolyases. The chromophores harness the energy of photoreactivating blue wavelengths of light and use them to catalyze the breakage of the pyrimidine dimer back to adjacent monomers. [Adapted fromFriedberg, E. C., Walker, G. C., Siede, W., Wood, R. D., Schultz, R. A., and Ellenberger, T. (Eds.). DNA Repair and Mutagenesis, 2nd ed., ASM, Washington, D.C., 2006.]...

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