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Institut de Biologie StructuraleGrenoble / France

How an electron and a proton modulate protein binding to DNA

The bacterial protein RsrR that coordinates a [2Fe-2S] cluster, regulates the expression of genes involved either directly or indirectly in cell redox equilibrium. The redox state of this iron-sulfur cluster controls RsrR binding to its site in DNA ; only the +2 oxidized form binds to the nucleic acid. In 2019, the IBS/Metallo group in collaboration with Pr. Nick Le Brun (University of East Anglia, UK) published the crystal structure of RsrR (Volbeda et al., JACS 2019) that showed an unprecedented coordination of its iron-sulfur cluster by Cys, Cys, Glu and His. The comparison of the oxidized and reduced protein structures revealed two conformations for a conserved tryptophan residue (W), called Out (in green in the figure) and In (in orange), and the concomitant displacement of a histidine (H).
In this new study, both groups, in collaboration with Dr Jean-Marie Mouesca (CEA-DRF-IRIG-DIESE-SyMMES-CAMPE), have solved the crystal structure of an RsrR-DNA complex where the three key resides (W, H and Y in black in the figure) adopt a conformation closer to the Out form. By combining chemical modification of W, site-directed mutagenesis, X-ray crystallography, quantum calculations, molecular dynamics and metadynamics simulations, they have shown that Out and In correspond to the oxidized and reduced forms, respectively. In addition, they have determined that its reduction (1 in the figure) changes the pKa of the H histidine towards more basic values ; the resulting doubly-protonated H (2) leads to the significant displacement of W, H and Y (3) ; the dipole moment of W responds to electrostatic changes caused by the reduction. Finally, the In form, thus produced, dissociates from DNA (4).
This study paves the way for the understanding of the mechanism by which the RsrR protein regulates the universal NAD cofactor synthesis and function. This cofactor is involved in photosynthesis, ATP production and cell respiration.

Electron and Proton Transfers Modulate DNA Binding by the Transcription Regulator RsrR. Crack JC, Amara P, Volbeda A, Mouesca JM, Rohac R, Pellicer Martinez MT, Huang CY, Gigarel O, Rinaldi C, Le Brun NE, Fontecilla-Camps JC. J Am Chem Soc, 142(11):5104-5116 (2020).