Very Fast Product Release and Catalytic Turnover of DNA Photolyase

Espagne, Agathe; Byrdin, Martin; Eker, A.P.M.; Brettel, Klaus

doi:10.1002/cbic.200900328

Cited by 17 publications

(14 citation statements)

References 24 publications

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“…[43] It is now well accepted that the CPD lesion features, in general, a relatively fast opening rate [44] (1.7 10 9 s À1 , [45] 1.8 10 9 s À1 , [46] and > 2 10 7 s À1 [42] ). The dimer cleavage efficiency in our competition experiment also allows us to estimate the kinetics of the rate-determining steps of CPD lesion cleavage in the duplex.…”

mentioning

confidence: 99%

Investigation of Excess‐Electron Transfer in DNA Double‐Duplex Systems Allows Estimation of Absolute Excess‐Electron Transfer and CPD Cleavage Rates

Fazio

Trindler

Heil

et al. 2010

Chemistry A European J

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To investigate the parameters and rates that determine excess-electron transfer processes in DNA duplexes, we developed a DNA double-duplex system containing a reduced and deprotonated flavin donor at the junction of two duplexes with either the same or different electron acceptors in the individual duplex substructures. This model system allows us to bring the two electron acceptors in the duplex substructures into direct competition for injected electrons and this enables us to decipher how the kind of acceptor influences the transfer data. Measurements with the electron acceptors 8-bromo-dA (BrdA), 8-bromo-dG (BrdG), 5-bromo-dU (BrdU), and a cyclobutane pyrimidine dimer, which is a UV-induced DNA lesion, allowed us to obtain directly the maximum overall reaction rates of these acceptors and especially of the T=T dimer with the injected electrons in the duplex. In line with previous observations, we detected that the overall dimer cleavage rate is about one order of magnitude slower than the debromination of BrdU. Furthermore, we present a more detailed explanation of why sequence dependence cannot be observed when a T=T dimer is used as the acceptor and we estimate the absolute excess-electron hopping rates.

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confidence: 99%

Investigation of Excess‐Electron Transfer in DNA Double‐Duplex Systems Allows Estimation of Absolute Excess‐Electron Transfer and CPD Cleavage Rates

Fazio

Trindler

Heil

et al. 2010

Chemistry A European J

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“…To our knowledge, the turnover rate for the repair by (6-4) PHR has not been reported. The turnover rate for the repair by CPD PHR was recently determined to be ~ 4 ms (at the substrate concentration of 250 μM) and the rate determing step of this rate was suggested to be the binding step 35 . This suggestion agrees with our above discussion.…”

Section: Discussionmentioning

confidence: 99%

Light-Induced Conformational Change and Product Release in DNA Repair by (6−4) Photolyase

et al. 2011

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Proteins of the cryptochrome/photolyase family share high sequence similarities, common folds and the flavin adenine dinucleotide (FAD) cofactor, but exhibit diverse physiological functions. Mammalian cryptochromes are essential regulatory components of the 24-hour circadian clock, whereas (6-4) photolyases recognize and repair UV-induced DNA damage by using light energy absorbed by FAD. Despite increasing knowledge about physiological functions from genetic analyses, the molecular mechanisms and conformational dynamics involved in clock signaling and DNA repair remain poorly understood. The (6-4) photolyase, which has strikingly high similarity to human clock cryptochromes, is a prototypic biological system to study conformational dynamics of cryptochrome/photolyase family proteins. The entire light-dependent DNA repair process for (6-4) photolyase can be reproduced in a simple in vitro system. To decipher pivotal reactions of the common FAD cofactor, we accomplished time resolved measurements of radical formation, diffusion, and protein conformational changes during light-dependent repair by full-length (6-4) photolyase on DNA carrying a single UV-induced damage. The (6-4) photolyase by itself showed significant volume changes after blue light activation, indicating protein conformational changes distant from the flavin cofactor. A drastic diffusion change was observed only in the presence of both (6-4) photolyase and damaged DNA, and not for (6-4) photolyase alone or with undamaged DNA. Thus, we propose that this diffusion change reflects the rapid (50 μs time constant) dissociation of the protein from the repaired DNA product. Conformational changes with such fast turnover would likely enable DNA repair photolyases to access the entire genome in cells.

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“…It is widely accepted that photoreactivation is an enzymatic process catalyzed by photolyase that follows the conventional Michaelis-Menten scheme [1,2,14]:…”

Section: Photoreactivation Of E Coli Cells Under Continuous Illuminamentioning

confidence: 99%

“…The reaction of photoreactivation follows a two-step scheme [1,2,14]: a light-independent step to form a complex between photolyase and a dimer lesion, followed by a light-dependent step to photorepair the lesion. The enzyme-substrate complex formation step is dependent on temperature, pH, and ionic strength, making the entire photoreactivation process to be influenced by these post-UV irradiation conditions [6,[15][16][17].…”

Section: Introductionmentioning

confidence: 99%