Institut de Biologie StructuraleGrenoble / France

Contact person(s) related to this article / TIMMINS Joanna

DNA damage recognition and repair

Our team has a long-standing interest in the DNA repair machinery of D. radiodurans, which is known to contribute to the survival of D. radiodurans after exposure to high doses of UV or ionizing radiation. D. radiodurans possesses a relatively ‘classical’ DNA repair repertoire, but, we and others, have shown that the small changes in the structure, substrate specificity and/or catalytic activities of these proteins have made them significantly more effective than their counterparts from radio-sensitive bacteria (Timmins and Moe, 2016).

We are particularly interested in the early steps of DNA repair during which the repair machinery detects and eliminates DNA lesions within a vast excess of undamaged DNA. This process is comparable to looking for a needle in a haystack ! How does a repair protein detect rare small base modifications, which do not necessarily alter the DNA duplex structure or the base-pairing, within millions of intact bases ?

To address this central question, our research focuses on the base excision (BER) and nucleotide excision (NER) repair pathways, which together are responsible for eliminating all damages occurring at the nucleotide level. We perform structural and functional studies of the major proteins involved in the initiation of these pathways, i.e. the DNA glycosylases in the case of BER and the UvrABC proteins for the NER pathway. Because DNA damage recognition relies on transient protein-DNA and protein-protein interactions, we are particularly interested in the dynamics of these processes and thus combine structural and biochemical studies with molecular dynamics simulations.

Members of the team

• Salvatore DE BONIS
• Joanna TIMMINS

Techniques

• Molecular biology
• Recombinant expression and purification of proteins
• Biochemical & biophysical characterisation of protein-DNA complexes
• Macromolecular crystallography
• DNA repair assays
• Lesion detection and quantification

Collaborations

• Jean-Luc RAVANAT (CEA/SyMMES, Grenoble)
• François DEHEZ & Antonio MONARI (LPCT, Nancy)
• Elin MOE (ITQB, Lisbon, Portugal)

Major publications

Sarre A, Stelter M, Rollo F, De Bonis S, Seck A, Hognon C, Ravanat JL, Monari A, Dehez F, Moe E and Timmins J. The three Endonuclease III variants of Deinococcus radiodurans possess distinct and complementary DNA repair activities. DNA Repair (2019) 78 p. 45-59. DOI : 10.1016/j.dnarep.2019.03.014.

Timmins J and Moe E. A decade of biochemical and structural studies of the DNA repair machinery of Deinococcus radiodurans. Review article. Comput Struct Biotechnol J. (2016) 14 p. 168-176. DOI : 10.1016/j.csbj.2016.04.001.

Sarre A, Ökvist M, Klar T, Hall DR, Smålas A, McSweeney S, Moe E and Timmins J. Structural and functional characterization of two unusual endonuclease III enzymes from Deinococcus radiodurans. Journal of Structural Biology (2015) 191 (2) p. 87-99. DOI : 10.1016/j.jsb.2015.05.009.

Stelter M, Acajjaoui S, McSweeney S and Timmins J. Structural and functional characterization of drUvrD provide new insights into DNA unwinding and helicase polarity. PLoS ONE (2013) 8 (10) : e77364. DOI : 10.1371/journal.pone.0077364.

Radzimanowski J, Dehez F, Round A, Bidon-Chanal A, McSweeney S and Timmins J. An ‘open’ structure of the RecOR complex supports ssDNA binding within the core of the complex. Nucleic Acids Research (2013) 41 (16) p.7972-7986. DOI : 10.1093/nar/gkt572.

Pellegrino S, Radzimanowski J, de Sanctis D, Boeri Erba E, McSweeney S and Timmins J. Structural and functional characterization of an SMC-like protein RecN : New insight into double-strand break repair. Structure (2012) 20 p.2076-2089. DOI : 10.1016/j.str.2012.09.010.

Moe E, Hall DR, Leiros I, Talstad V, Timmins J, McSweeney S. Structure/function studies of an unusual 3-methyladenine DNA glycosylase II (AlkA) from Deinococcus radiodurans. Acta Crys. (2012) D68 p.703-712. DOI : 10.1107/S090744491200947X.

Timmins J, Gordon E, Caria S, Leonard G, Acajjaoui S, Kuo MS, Monchois V, McSweeney S. Structural and mutational analyses of Deinococcus radiodurans UvrA2 provide insight into DNA binding and damage recognition by UvrAs. Structure. (2009) 17 (4) p.547-58. DOI : 10.1016/j.str.2009.02.008.

Timmins J, Leiros I, McSweeney S. Crystal structure and mutational study of RecOR provide insight into its mode of DNA binding. EMBO J. (2007) 26 (13) p.3260-71. DOI : 10.1038/sj.emboj.7601760.

Timmins J, Leiros I, Hall DR, McSweeney S. Crystal structure and DNA-binding analysis of RecO from Deinococcus radiodurans. EMBO J. (2005) 24 (5) p.906-18. DOI : 10.1038/sj.emboj.7600582.