The Lrp/AsnC family of transcriptional regulators is commonly found in prokaryotes and is associated with the regulation of amino acid metabolism. However, it remains unclear how the L-cysteine-responsive Lrp/AsnC family regulator perceives and responds to L-cysteine. Here, we try to elucidate the molecular mechanism of the L-cysteine-responsive transcriptional regulator. Through 5'RACE and EMSA, we discovered a 15 bp incompletely complementary pair palindromic sequence essential for DecR binding, which differed slightly from the binding sequence of other Lrp/AsnC transcription regulators. Using alanine scanning, we identified the L-cysteine binding site on DecR and found that different Lrp/AsnC regulators adjust their binding pocket's side-chain residues to accommodate their specific effector. MD simulations were then conducted to explore how ligand binding influences the allosteric behavior of the protein. PCA and in silico docking revealed that ligand binding induced perturbations in the linker region, triggering conformational alterations and leading to the relocalization of the DNA-binding domains, enabling the embedding of the DNA-binding region of DecR into the DNA molecule, thereby enhancing DNA-binding affinity. Our findings can broaden the understanding of the recognition and regulatory mechanisms of the Lrp/AsnC-type transcription factors, providing a theoretical basis for further investigating the molecular mechanisms of other transcription factors.
Keywords: Allosteric regulation; Cis-acting motif; DecR; L-cysteine binding region; Transcriptional regulator.
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