Abstract
YfdX is a prokaryotic protein encoded by several pathogenic bacteria including Salmonella enterica serovar Typhi, which causes one of the most fatal infectious diseases, typhoid fever. YfdX is a product of the yfdXWUVE operon and is known to be under the control of EvgA, a regulator protein controlling the expression of several proteins involved in response to environmental stress, in Escherichia coli. Nevertheless, unlike other proteins encoded by the same operon, the structural and physiological aspects of YfdX have been poorly characterized. Here, we identified a previously unknown pH-dependent stoichiometric conversion of S. Typhi YfdX between dimeric and tetrameric states; this conversion was further analyzed via determining its structure by X-ray crystallography at high resolution and by small-angle X-ray scattering in a solution state and via structure-based mutant studies. Biologically, YfdX was proven to be critically involved in Salmonella susceptibility to two β-lactam antibiotics, penicillin G and carbenicillin, as bacterial growth significantly impaired by its deficiency upon treatment with each of the two antibiotics was recovered by chromosomal complementation. Furthermore, by using Galleria mellonella larvae as an in vivo model of Salmonella infection, we demonstrated that Salmonella virulence was remarkably enhanced by YfdX deficiency, which was complemented by a transient expression of the wild-type or dimeric mutant but not by that of the monomeric mutant. The present study work provides direct evidence regarding the participation of YfdX in Salmonella antibiotic susceptibility and in the modulation of bacterial virulence, providing a new insight into this pathogen’s strategies for survival and growth.
Highlights
Salmonella enterica serovar Typhi is a gram-negative bacterium that infects humans only (Hurley et al, 2014), causing systemic typhoid fever
By using Galleria mellonella larvae as an in vivo model of Salmonella infection, we demonstrated that Salmonella virulence was remarkably enhanced by YfdX deficiency, which was complemented by a transient expression of the wild-type or dimeric mutant but not by that of the monomeric mutant
Typhi YfdX protein containing residues 10–186 from its total 199 amino acids, referred to as st_YfdX in this manuscript, was overexpressed in E. coli and purified on a Ni–NTA column and a size-exclusion chromatography column, equilibrated with a final buffer consisting of 20 mM Tris-HCl, 200 mM NaCl, and 1 mM DTT (Figure 1A)
Summary
Salmonella enterica serovar Typhi is a gram-negative bacterium that infects humans only (Hurley et al, 2014), causing systemic typhoid fever. S. Typhi initially penetrates the small intestinal epithelial cells and spreads through the bloodstream to other organs such as the spleen, liver, and bone marrow, where this bacterium multiplies and reenters the bloodstream causing symptoms including a high fever (Everest et al, 2001). Typhi to acquire resistance to various antibiotics through genetic changes: a phenomenon called “multiple drug resistance (MDR)” (Rowe et al, 1997; Kalra et al, 2003; Frye and Jackson, 2013). MDR strains exhibit strong resistance to antibiotics, and lead to treatment failure (Rowe et al, 1997; Parkhill et al, 2001; Kariuki et al, 2015; Karkey et al, 2018)
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