9CA6

Rns Pocket Mutant - I14F/I17F

Summary for 9CA6

• Entry DOI: 10.2210/pdb9ca6/pdb
• Descriptor: Regulatory protein Rns (1 entity in total)
• Functional Keywords: arac family, etec, dna binding protein
• Biological source: Escherichia coli
• Total number of polymer chains: 2
• Total formula weight: 61759.64

Authors

Tolbert, J.D.,Midgett, C.R.,Kull, F.J. (deposition date: 2024-06-17, release date: 2025-06-25, Last modification date: 2026-08-05)

Primary citation

Tolbert, J.D.,Talbot, K.M.,Bollinger, C.M.,Kull, F.J.,Munson, G.P.,Midgett, C.R.
Characterization of the ligand binding pocket of the virulence regulator Rns, a member of the AraC/XylS family of transcription factors.
Msphere, 10:e0011525-e0011525, 2025

PubMed Abstract: Diarrheal disease caused by Gram-negative enteric pathogens, such as enterotoxigenic (ETEC), , spp., and spp., is a leading cause of morbidity and mortality of children, especially in low resource nations. While progress has been made in reducing this burden, there remains a need to develop effective therapies. Recently, we determined the structure of Rns, a member of the AraC/XylS family that regulates the expression of pili and other virulence factors in ETEC. The structure revealed decanoic acid bound between the N- and C-terminal domains. To test the hypothesis that bound decanoic acid directly inhibits Rns, we identified amino acid side chains predicted to be necessary for ligand binding. Removal of the positive side chains of R75 and H20 rendered Rns insensitive to fatty acid inhibition. Additionally, mutations designed to block decanoic acid binding also produced a variant Rns that was fatty acid insensitive. We also observed that this variant is structurally more flexible than wildtype Rns bound to decanoic acid, suggesting that fatty acid binding contributes to structural rigidity. These studies demonstrate that Rns binding pocket residues are critical for binding fatty acids, which result in inhibition of DNA binding and support our hypothesis that fatty acids must bind in the binding pocket to inhibit other AraC regulators. Further work by us and others suggests that inhibition of AraC virulence regulators by fatty acids is a common paradigm among many bacterial pathogens. Therefore, understanding the molecular basis of inhibition lays the groundwork for the development of small molecule therapeutics targeting enteric disease.

PubMed: 40704798
DOI: 10.1128/msphere.00115-25

Experimental method

X-RAY DIFFRACTION (3 Å)