Structural basis of the conformational changes in Microbacterium hydrocarbonoxydans IclR transcription factor homolog due to ligand binding

Functional and Structural Characterization of an IclR Family Transcription Factor for the Development of Dicarboxylic Acid Biosensors

“…The homodimer in the ligandbound form retains an overall structure similar to that of the apo structure but undergoes key localized ligand-induced conformational changes. Similarly to the mechanism proposed for pHbrR, the exterior opening of the binding pocket in the PcaR/succinate complex closes upon the ligand's binding (Figures 3C and 3D) [23]. Like other IclR family members including E. coli IclR, the binding pocket is centrally located and formed by six-stranded anti-parallel β-sheets and two long α-helixes (corresponding to α6 and α10 in PcaR) on one side and three (α7 to α9) short α-helixes on the other side (Figure 5A).…”

“…Out of such attempts, a succinate-bound complex structure was solved in the same space group as the apo structure, and the homodimer was again present in the asymmetric unit. This succinate-bound complex complements the apo structure as only the second pair of full-length crystal structures of an IclR family representative in both ligand-bound and unliganded form, with the only other one being from pHbrR [23]. Analysis of the ligand-bound structure reveals that PcaR accommodates succinate in a binding pocket of the C-terminal domain on both subunits of protein's dimer (Figures 3D and 5A).…”

“…BaaR [41], BlcR [42], and pHbrR [23], the PcaR structure contains two polypeptide molecules in the asymmetric unit, reflecting the dimeric nature of this TF in solution. Following the IclR family's characteristic architecture [5], each PcaR molecule consists of two domains, the Nterminal winged HTH DBD, and C-terminal LBD, with an α-helix linker region connecting these two domains (Figure 3A).…”

“…Currently there are only five representatives of the IclR the family that have been structurally characterized in complex with their corresponding ligands, including: E. coli IclR in complex with pyruvate (PDB: 2O9A) or glyoxylate (PDB: 2O99) [13]; PobR with parahydroxybenzoate (PDB: 5W1E) [22]; a PobR mutant with altered specificity bound to 3hydroxybenzoate (PDB: 5HPI) [9]; and most recently pHbrR bound to 4-hydroxybenzoate (PDB: 7CUO) [23]. The structure of BaaR in complex with acetate in the binding pocket has also been determined (PDB: 5WHM), but this acetate molecule derives from the crystallization buffer [24].…”

“…The structure of BaaR in complex with acetate in the binding pocket has also been determined (PDB: 5WHM), but this acetate molecule derives from the crystallization buffer [24]. Previous work has shown that BaaR maintains an open loop formation when binding to acetate in the binding pocket, unlike what is seen in other examples like pHbrR where binding leads to a closed loop, indicating that acetate is likely not a ligand of BaaR [23]. Such a low number of IclR-ligand complexes reflects the limited understanding of the basis of ligand recognition by IclRs and hinders rational design of IclR-based TFs as biosensors.…”