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Molecular movie of DNA repair by a photolyase

​​​​An international collaboration involving researchers at IRIG has revealed the details of DNA lesion repair by the enzyme photolyase using time-resolved femtosecond crystallography, from lesion bond breaking to enzyme/DNA complex dissociation.​
Published on 4 December 2023

DNA damage induced by ultraviolet radiation can be fixed in many organisms by a light-activated enzyme known as photolyase. Researchers at IRIG have been involved in studying the mechanism of a methanogenic archaea photolyase that specifically repairs cyclobutane pyrimidine dimer (CPD) lesions. Using visible light, photolyase catalyzes the rupture of two covalent bonds linking two adjacent pyrimidine rings in a DNA strand. ​​
The study of the molecular mechanism of DNA strand repair was carried out at the free-electron X-ray lasers (XFEL) SwissFEL in Switzerland and SACLA in Japan using time-resolved crystallography. Researchers at IRIG have analyzed spectroscopic and crystallographic data, on a timescale between 100 picoseconds and 200 microseconds covering more than 6 orders of magnitude in order to reveal an unprecedented level of detail. The mechanism consists in the transfer of one electron from the FAD (Flavine Adenine Dinucleotide) cofactor to the damaged DNA, the sequential breaking of two covalent bonds, the rearrangement of the various chemical groups involved in the reaction, and, finally, the back-flipping of the two repaired DNA bases, which leads to the dissociation of the enzyme/DNA complex. (cf. Figure). ​​

​​Figure: On the left: the damaged bases of the DNA strand point towards the inside of the enzyme photolyase. Upon blue light absorption by the FAD cofactor, the covalent bonds of the CPD lesion are sequentially broken in less than 10 nanoseconds, as illustrated by the difference electron density map (in orange and blue in the upper inset). On the right: after 200 microseconds, the repaired bases have reoriented towards the inside of the DNA double-strand, which is then ready to dissociate from the enzyme. ​
​​© Nicolas Caramello (ESRF and University of Hamburg) and Antoine Royant (IBS)

The data collected in this study constitutes a genuine molecular movie of all events, from the absorption of a blue photon by the cofactor to the release of the fully repaired DNA double-strand.

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