6-4 Photolyase E. coli

Grystal structures of photo lyases from E. coli (Park ei al., 1995), A. nidulans (Tamada et al, 1997), and Thermus thermophilus (Komori et al, 2001) have been solved. Although they possess different second chromo-phores E. coli photolyase has MTHF and the latter two contain 8-HDF), and the level of overall sequence homology among the three enzymes is only about 25% sequence identity, the structures of all three are remarkably similar. Here, the structure of E. coli photolyase will be presented and the minor differences of the other two photolyases from this structure will be briefly mentioned. 

Brash, D. E., Franklin, W. A., Sancar, G. B., Sancar, A., and Haseltine, W. A. (1985). E. coli photolyase reverses cyclobutane pyrimidine dimers but not pyrimidine-pyrimidone (6-4) photoproducts. / Biol. Chem. 260, 11438-11441. 

Kim, S. T., Heelis, P. F., Okamura, T., Hirata, Y, Mataga, N., and Sancar, A. (1991). Determination of rates and yields of interchromophore enei transfer and inter-molecular electron transfer in E. coli photolyase by time-resolved fluorescence and absorption spectroscopy. Biochemistry 30, 11262—11270. 

Figure 23-49 Overall view of the DNA photolyase structure from E. coli. The ribbon traces the 471-residue chain. The bound cofactors FAD (left) and 5,10-methenyltetrahy-drofolate (right) are shown in ball-and-stick representation. From Park et al.652 Courtesy of Johan Deisenhofer.

An example of direct repair is the photochemical cleavage of pyrimidine dimers. Nearly all cells contain a photoreactivating enzyme called DNA photolyase. The E. coli enzyme, a 35-kd protein that contains bound N lO-methenyltetrahydrofolate and flavin adenine dinucleotide cofactors, binds to the distorted region of DNA. The enzyme uses light energy—specifically, the absorption of a photon by the N, N lO-methenyltetrahydrofolate coenzyme—to form an excited state that cleaves the dimer into its original bases. 

Whitfield, J. Morelli, A. Warner, J. C. Enzymatic Reversal of Polymeric Thymine Photocrosslinking with E. coli DNA Photolyase. J. Macromol. Sci. 2005,442, 1541-1546. 

An example of direct repair is the photochemical cleavage of pyrimidine dimers. Nearly all cells contain a photoreactivating enzyme called DjV4 photolyase. The E, coli enzyme, a 35-kd protein that contains bound N ... 

To elucidate further the possible role of flavin as a sensitizer in photolyase catalysis, Joms conducted a study on the light-catalyzed monomerization of thymine dimers by reduced flavins and flavin analogs as models for the DNA-PL reaction of E. coli and yeast (162). The reduced forms of 1-deazariboflavin, A(3)-methyllumiflavin, 7,8-dimethyl-1,10-ethyleneisoalloxazinium perchlorate, and 5-deazariboflavin were generated anaerobically with excess dithionite. Of these four flavins, cleavage of cij-syn-[me//iy/- H]thymine dimer to [methyl- H]-thymine was observed with reduced 1-deazariboflavin, and to a lesser extent with... 

Fig. 2 Pulsed ENDOR on a neutral flavin radical. E. coli CPD photolyase was investigated with pulsed Davies ENDOR spectroscopy at r = 80 K (for details, see [25]). Detectable protons are marked accordingly...

Fig. 7. Absorption and action spectra of DNA photolyases. Left, Escherichia coli photolyase. Solid and broken lines represent the absorption spectra of the E-MTHF-FADH and the E-FADH forms of the enzyme, and the triangles and squares represent the photolytic cross sections (exO) of the two forms. Right, Anacystis nidulans photolyase. The solid and broken lines are the absorption spectra of the E-8-HDF-FADH and the E-FADH forms of the enzyme, and the circles and triangles represent photolytic cross sections of the corresponding forms at selected wavelengths.

Husain, I., and Sancar, A. (1987). Binding of E. coli DNA photolyase to defined substrate containing a single ToT dimer. Nucleic Adds Bes. 15, 1109—1120. 

Joms, M. S., Sancar, G. B., and Sancar, A. (1984). Identification of a neutral flavin radical and characterization of a second chromophore in E. coli DNA photolyase. Biochemistry 23, 2673-2679. 

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