Structural insights into the DNA recognition mechanism by the bacterial transcription factor PdxR

This is the final version. Available from Oxford University Press via the DOI in this record.Atomic coordinates and structure factors for the reported apo-PdxR crystal structure have been deposited with the RCSB Protein Data Bank (PDB) under accession number 7PQ9. The cryo-EM maps of the holo-PdxR–DNA complex in the open, half-closed, and closed (C1 and C2 symmetry) conformation and the relative coordinates generated and analysed in the current study have been deposited in the Electron Microscopy Data Bank (EMDB) and in the PDB under accession code EMD-14960 (PDB 7ZTH), EMD-14778 (PDB 7ZLA), EMD-14852 (PDB 7ZPA) and EMD-14801 (PDB 7ZN5), respectively.Specificity in protein-DNA recognition arises from the synergy of several factors that stem from the structural and chemical signatures encoded within the targeted DNA molecule. Here, we deciphered the nature of the interactions driving DNA recognition and binding by the bacterial transcription factor PdxR, a member of the MocR family responsible for the regulation of pyridoxal 5'-phosphate (PLP) biosynthesis. Single particle cryo-EM performed on the PLP-PdxR bound to its target DNA enabled the isolation of three conformers of the complex, which may be considered as snapshots of the binding process. Moreover, the resolution of an apo-PdxR crystallographic structure provided a detailed description of the transition of the effector domain to the holo-PdxR form triggered by the binding of the PLP effector molecule. Binding analyses of mutated DNA sequences using both wild type and PdxR variants revealed a central role of electrostatic interactions and of the intrinsic asymmetric bending of the DNA in allosterically guiding the holo-PdxR-DNA recognition process, from the first encounter through the fully bound state. Our results detail the structure and dynamics of the PdxR-DNA complex, clarifying the mechanism governing the DNA-binding mode of the holo-PdxR and the regulation features of the MocR family of transcription factors.Italian MIUR-PRIN 2020POR FESR Lazio 2014–2020Sapienza University of RomeDefence Science and Technology Laboratory (DSTL)Istituto Pasteur Italia – Fondazione Cenci Bolognett