Biophysical and Functional Characterization of the Persulfide Sensor CstR from Staphylococcus aureus

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Date
2015-04
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[Bloomington, Ind.] : Indiana University
Abstract
How cells regulate the intracellular bioavailable pool of sulfur while mitigating its toxic effects is poorly understood. Major components of this pool, collectively referred to as reactive sulfur species (RSS), include hydrogen sulfide (H2S) and low molecular weight per- and polysulfides and become deleterious at increasing concentrations. For example, H2S functions as a gaseous signaling molecule but at elevated levels can poison cytochrome c oxidase of the electron transport chain. The human pathogen Staphylococcus aureus contains an apparent complete sulfur oxidation system, termed the cst operon, which is under control of the transcriptional regulator CstR (Copper-sensing operon repressor (CsoR)-like sulfurtransferase repressor). In this work, sulfide stress, as exogenous H2S or RSS, is identified as a strong inducer of the cst operon. Failure to express this operon or deletions of enzymes within results in loss cellular viability under sulfide stress conditions. Biophysical investigations into the regualtor CstR revealed it does not react with hydrogen sulfide directly but rather reacts with the sulfane sulfur of an inorganic polysulfide or organic persulfide donor to form di-, tri, and tetrasulfide bonds between two conserved cysteine residues, Cys31 and Cys60' on opposite protomers as determined by high-resolution tandem mass spectrometry. These modifications result in negative regulation of cst operator DNA binding affinity. Interestingly, CstR displays reaction specificity compared to the structurally similar CsoR that is also found in S. aureus, which controls the expression of genes responsible for copper toxicity resistance. Comparative analysis of CstR and CsoR cysteine reactivity by pulsed-alkylation mass spectrometry reveals a striking difference in Cys31 reactivity (Cys41 in CsoR). These studies provide the framework for better understanding of sulfide homeostasis and stress resistance as well as identification of key differences in CstR and CsoR inducer selectivity.
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Thesis (Ph.D.) - Indiana University, Chemistry, 2015
Keywords
CstR, Hydrogen Sulfide, Oxidative Stress, Persulfide, Staphylococcus aureus, Sulfur Metabolism
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Doctoral Dissertation