Sulisobenzone is a potent inhibitor of the global transcription factor Cra

Neetu Neetu; Jai Krishna Mahto; Monica Sharma; Madhusudhanarao Katiki; Preeti Dhaka; Partha Roy; Shailly Tomar; Anoop Narayan; Dinesh Yernool; Pravindra Kumar

doi:10.1016/j.jsb.2023.108034

Sulisobenzone is a potent inhibitor of the global transcription factor Cra

J Struct Biol. 2023 Dec;215(4):108034. doi: 10.1016/j.jsb.2023.108034. Epub 2023 Oct 5.

Authors

Affiliations

¹ Department of Biosciences and Bioengineering, Indian Institute of Technology Roorkee, Roorkee, Uttarakhand, India.
² Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA.
³ Department of Biological Sciences, Purdue University, West Lafayette, IN 47906, USA.
⁴ Department of Biosciences and Bioengineering, Indian Institute of Technology Roorkee, Roorkee, Uttarakhand, India. Electronic address: pravindra.kumar@bt.iitr.ac.in.

PMID: 37805153
DOI: 10.1016/j.jsb.2023.108034

Abstract

Transcription is carried out by the RNA polymerase and is regulated through a series of interactions with transcription factors. Catabolite activator repressor (Cra), a LacI family transcription factor regulates the virulence gene expression in Enterohaemorrhagic Escherichia coli (EHEC) and thus is a promising drug target for the discovery of antivirulence molecules. Here, we report the crystal structure of the effector molecule binding domain of Cra from E. coli (EcCra) in complex with HEPES molecule. Based on the EcCra-HEPES complex structure, ligand screening was performed that identified sulisobenzone as an potential inhibitor of EcCra. The electrophoretic mobility shift assay (EMSA) and in vitro transcription assay validated the sulisobenzone binding to EcCra. Moreover, the isothermal titration calorimetry (ITC) experiments demonstrated a 40-fold higher binding affinity of sulisobenzone (K_D 360 nM) compared to the HEPES molecule. Finally, the sulisobenzone bound EcCra complex crystal structure was determined to elucidate the binding mechanism of sulisobenzone to the effector binding pocket of EcCra. Together, this study suggests that sulisobenzone may be a promising candidate that can be studied and developed as an effective antivirulence agent against EHEC.

Keywords: Drug discovery; Electrophoretic mobility shift assay; In vitro transcription assay; Isothermal titration calorimetry; Protein crystallization; Sulisobenzone; Transcription factor; X-ray crystallography.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Escherichia coli* / metabolism
Gene Expression Regulation, Bacterial
HEPES / metabolism
Protein Binding
Repressor Proteins / genetics
Transcription Factors* / metabolism

Substances

Transcription Factors
sulisobenzone
Repressor Proteins
HEPES