Binding of Pyrimidine Model Dimers to the Photolyase Enzyme:  A Molecular Dynamics Study

Hahn, Jutta; Michel‐Beyerle, M. E.; Rösch, Notker

doi:10.1021/jp984197h

Cited by 30 publications

(41 citation statements)

References 26 publications

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“…The data support the hypothesis that the binding geometry within the active site and not the substitution pattern of the cyclobutane moiety determines the higher enzymatic repair rate of uridine type DNA lesions. This result is in good agreement with recent ultrashort time spectroscopic data of Michel-Beyerle [18] and calculation of Rösch [17] , which suggest that the two thymidine methyl groups hinder the thymidine lesion to penetrate as deep as the uridine substrate into the active site. [18] Calculations: In order to investigate how the structure of the dimer influences the cleavage rate ab initio calculations were performed [11] with the X-ray crystal structural data of 12 and 13 as input geometries for the uracil and thymine dimers in their neutral and radical anion states.…”

supporting

confidence: 91%

“…Different binding geometries could determine the repair rate by influencing the electron transfer processes between the FADH À cofactor and the corresponding lesion. [17,18] In order to investigate the molecular reasons for the different repair efficiencies in detail, we studied the cleavage rates of uracil and thymine dimers inside and outside the photolyase active site. To this end, the two model compounds 1 and 2 (Scheme 1) were prepared and investigated.…”

Section: Introductionmentioning

confidence: 99%

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A Comparative Repair Study of Thymine- and Uracil-Photodimers with Model Compounds and a Photolyase Repair Enzyme

et al. 2000

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Cyclobutane uridine and thymidine dimers with cis-syn-structure are DNA lesions, which are efficiently repaired in many species by DNA photolyases. The essential step of the repair reaction is a light driven electron transfer from a reduced FAD cofactor (FADH ) to the dimer lesion, which splits spontaneously into the monomers. Repair studies with UV-light damaged DNA revealed significant rate differences for the various dimer lesions. In particular the effect of the almost eclipsed positioned methyl groups at the thymidine cyclobutane dimer moiety on the splitting rates is unknown. In order to investigate the cleavage vulnerability of thymine and uracil cyclobutane photodimers outside the protein environment, two model compounds, containing a thymine or a uracil dimer and a covalently connected flavin, were prepared and comparatively investigated. Cleavage investigations under internal competition conditions revealed, in contrast to all previous findings, faster repair of the sterically less encumbered uracil dimer. Stereoelectronic effects are offered as a possible explanation. Ab initio calculations and X-ray crystal structure data reveal a different cyclobutane ring pucker of the uracil dimer, which leads to a better overlap of the pi*-C(4)-O(4)-orbital with the sigma*-C(5)-C(5')-orbital. Enzymatic studies with a DNA photolyase (A. nidulans) and oligonucleotides, which contain either a uridine or a thymidine dimer analogue, showed comparable repair efficiencies for both dimer lesions. Under internal competition conditions significantly faster repair of uridine dimers is observed.

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confidence: 91%

Section: Introductionmentioning

confidence: 99%

A Comparative Repair Study of Thymine- and Uracil-Photodimers with Model Compounds and a Photolyase Repair Enzyme

et al. 2000

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“…6). The protein is bound to the DNA oligomer at the location of the dimer lesion, and the dimer lies inside the protein's active site [a binding mode also suggested by biochemical, fluorescence, and NMR experiments (1)(2)(3)(7)(8)(9), and investigated in several computational studies (10)(11)(12)(13)(14)]. In the active site, the dimer is adjacent to FADH Ϫ (6).…”

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confidence: 99%

Photoselected electron transfer pathways in DNA photolyase

Prytkova

Beratan

Skourtis

2007

Proc. Natl. Acad. Sci. U.S.A.

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Cyclobutane dimer photolyases are proteins that bind to UVdamaged DNA containing cyclobutane pyrimidine dimer lesions. They repair these lesions by photo-induced electron transfer. The electron donor cofactor of a photolyase is a two-electron-reduced flavin adenine dinucleotide (FADH ؊ ). When FADH ؊ is photo-excited, it transfers an electron from an excited 3 * singlet state to the pyrimidine dimer lesion of DNA. We compute the lowest excited singlet states of FADH ؊ using ab initio (time-dependent density functional theory and time-dependent Hartree-Fock), and semiempirical (INDO͞S configuration interaction) methods. The calculations show that the two lowest 3 * singlet states of FADH ؊ are localized on the side of the flavin ring that is proximal to the dimer lesion of DNA. For the lowest-energy donor excited state of FADH ؊ , we compute the conformationally averaged electronic coupling to acceptor states of the thymine dimer. The coupling calculations are performed at the INDO͞S level, on donor-acceptor cofactor conformations obtained from molecular dynamics simulations of the solvated protein with a thymine dimer docked in its active site. These calculations demonstrate that the localization of the 1 FADH ؊ * donor state on the flavin ring enhances the electronic coupling between the flavin and the dimer by permitting shorter electron-transfer pathways to the dimer that have single through-space jumps. Therefore, in photolyase, the photo-excitation itself enhances the electron transfer rate by moving the electron towards the dimer.

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“…Alanine substitution mutagenesis studies by Berg and Sancar (10) also support this hypothesis. The question has sparked several computational studies as well, most of which support the base flipping hypothesis (11,12). Thermodynamic studies suggest that there is little energy cost for flipping the CPD into solution (13).…”

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confidence: 99%

Cyclobutylpyrimidine Dimer Base Flipping by DNA Photolyase

Christine

Macfarlane

Yang

et al. 2002

Journal of Biological Chemistry

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DNA Photolyase is a flavoprotein that uses light to repair cyclobutylpyrimidine dimers in DNA. From considerations of the crystal structure of the protein, it has been hypothesized that the dimer lesion is flipped out of the DNA double helix into the substrate binding pocket. We have used a fluorescent adenine analog, 2-aminopurine (2-Ap), as a probe of local double helical structure upon binding of the substrate to the protein. Our results show that the local structure around the thymidine lesion changes dramatically upon binding to Photolyase. This is consistent with base flipping of the lesion into the protein binding cavity with concomitant destacking of the opposing complementary 2-Ap nucleotide.Cyclobutylpyrimidine dimers (CPDs) 1 are lesions formed in DNA between adjacent pyrimidines upon absorption of UV light. These lesions cause replicational errors and can lead to cell death or cancer if left unrepaired (1, 2). One repair protein, DNA photolyase, incorporates a non-covalently bound FAD and requires blue light for repair (3). There is ample evidence that repair of the CPD proceeds by electron transfer from a photoexcited fully reduced FADH Ϫ to the CPD, which subsequently monomerizes within 2 ns (3). Studies by Payne et al. (4) have demonstrated that the oxidized enzyme can bind CPD-containing DNA but cannot efficiently repair the CPD lesion. As we show below, this differential behavior is extremely useful in understanding the substrate binding mode of photolyase.The crystal structures of the Escherichia coli and Aspergillus nidulans holoenzymes were solved by Kim et al. (5) and Tamada and co-workers (6), respectively. These crystal structures revealed important structural elements that were both familiar and surprising. It was noted that the cavity has approximately the correct dimensions to enclose the CPD if the thymidines were folded in a coplanar geometry, leading to the prediction that photolyase would bind the CPD by "flipping out" the lesion from the double helical DNA (5). The FADH Ϫ cofactor was found to lie at the bottom of this cavity, consistent with the ability of the cofactor to resist oxidation by molecular O 2 . The surface of the protein above the cavity incorporates a strip of positively charged amino acid residues that are thought to help orient the negatively charged phosphate backbone of the damaged DNA strand.Unfortunately, no crystal structure of the enzyme-substrate complex is available, but a recent crystal structure of Thermus thermophilus complexed with thymine was published by Komori and co-workers (7), which shows the thymine residing in the putative substrate cavity. While thymine is not a substrate, it can be thought of as a partial product of the repair reaction. This is the strongest evidence regarding the substrate binding mode of photolyase for CPDs. However, it does not clarify whether one or both of the thymine bases of the CPD is bound in the cavity.Other experimental evidence for the mode of substrate binding is available. Most notable are the ethylation and ...

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Binding of Pyrimidine Model Dimers to the Photolyase Enzyme: A Molecular Dynamics Study

Cited by 30 publications

References 26 publications

A Comparative Repair Study of Thymine- and Uracil-Photodimers with Model Compounds and a Photolyase Repair Enzyme

A Comparative Repair Study of Thymine- and Uracil-Photodimers with Model Compounds and a Photolyase Repair Enzyme

Photoselected electron transfer pathways in DNA photolyase

Cyclobutylpyrimidine Dimer Base Flipping by DNA Photolyase

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