Investigation of Zur-regulated metal transport systems reveals an unexpected role of pyochelin in zinc homeostasis.

Unlabelled: Limiting the availability of transition metals at infection sites serves as a critical defense mechanism employed by the innate immune system to combat microbial infections. exhibits a remarkable ability to thrive in zinc-deficient environments, facilitated by intricate cellular responses governed by numerous genes regulated by the zinc-responsive transcription factor Zur. Many of these genes have unknown functions, including those within the predicted and operons.

TRAP1 S-nitrosylation as a model of population-shift mechanism to study the effects of nitric oxide on redox-sensitive oncoproteins.

S-nitrosylation is a post-translational modification in which nitric oxide (NO) binds to the thiol group of cysteine, generating an S-nitrosothiol (SNO) adduct. S-nitrosylation has different physiological roles, and its alteration has also been linked to a growing list of pathologies, including cancer. SNO can affect the function and stability of different proteins, such as the mitochondrial chaperone TRAP1.

Localized Infections with Strains Defective in Zinc Uptake Reveal That Zebrafish Embryos Recapitulate Nutritional Immunity Responses of Higher Eukaryotes.

The innate immune responses of mammals to microbial infections include strategies based on manipulating the local concentration of metals such as iron (Fe) and zinc (Zn), commonly described as nutritional immunity. To evaluate whether these strategies are also present in zebrafish embryos, we have conducted a series of heart cavity-localized infection experiments with strains characterized by a different ability to acquire Zn. We have found that, 48 h after infection, the bacterial strains lacking critical components of the Zn importers ZnuABC and ZrmABCD have a reduced colonization capacity compared to the wild-type strain.

Zinc-binding metallophores protect Pseudomonas aeruginosa from calprotectin-mediated metal starvation.

Pseudomonas aeruginosa is known to exhibit considerable resistance to the antimicrobial activity of the metal-sequestering protein calprotectin (CP). In this study, we demonstrate that although CP induces zinc deficiency in P. aeruginosa, a strain unable to import zinc through the two most important metal acquisition systems, namely ZnuABC and ZrmABCD, maintains significant growth capacity in the presence of high concentrations of CP.

Typhimurium and Respond Differently to the Fe Chelator Deferiprone and to Some Novel Deferiprone Derivatives.

The ability to obtain Fe is critical for pathogens to multiply in their host. For this reason, there is significant interest in the identification of compounds that might interfere with Fe management in bacteria. Here we have tested the response of two Gram-negative pathogens, serovar Typhimurium (STM) and (PAO1), to deferiprone (DFP), a chelating agent already in use for the treatment of thalassemia, and to some DFP derivatives designed to increase its lipophilicity.

S-nitrosylation affects TRAP1 structure and ATPase activity and modulates cell response to apoptotic stimuli.

The mitochondrial chaperone TRAP1 has been involved in several mitochondrial functions, and modulation of its expression/activity has been suggested to play a role in the metabolic reprogramming distinctive of cancer cells. TRAP1 posttranslational modifications, i.e.

Growth of Pseudomonas aeruginosa in zinc poor environments is promoted by a nicotianamine-related metallophore.

Previous studies have suggested that P. aeruginosa possesses redundant zinc uptake systems. To identify uncharacterized zinc transporters, we analyzed the genome-wide transcriptional responses of P.

An ERp57-mediated disulphide exchange promotes the interaction between Burkholderia cenocepacia and epithelial respiratory cells.

Previous studies have demonstrated that extracellular glutathione reduces the ability of the Cystic Fibrosis pathogen Burkholderia cenocepacia to infect primary or immortalized epithelial respiratory cells. We report here that the adhesion and invasion ability of B. cenocepacia is limited also by thiol-oxidizing and disulphide-reducing agents and by protein disulfide isomerase (PDI) inhibitors.

The capability of Pseudomonas aeruginosa to recruit zinc under conditions of limited metal availability is affected by inactivation of the ZnuABC transporter.

The ability of a large number of bacterial pathogens to multiply in the infected host and cause disease is dependent on their ability to express high affinity zinc importers. In many bacteria, ZnuABC, a transporter of the ABC family, plays a central role in the process of zinc uptake in zinc poor environments, including the tissues of the infected host. To initiate an investigation into the relevance of the zinc uptake apparatus for Pseudomonas aeruginosa pathogenicity, we have generated a znuA mutant in the PA14 strain.

Extracellular glutathione decreases the ability of Burkholderia cenocepacia to penetrate into epithelial cells and to induce an inflammatory response.

Background: The airway surface liquid (ASL) of Cystic Fibrosis (CF) patients contains a lower concentration of reduced glutathione (GSH) with respect to healthy people. It is not known whether this defect may favor lung colonization by opportunistic pathogens.

Principal Findings: We have analyzed the effects of extracellular GSH on the ability of Burkholderia cenocepacia to penetrate and multiply in epithelial respiratory cells.

Low-Shear Modeled Microgravity Enhances Salmonella Enterica Resistance to Hydrogen Peroxide Through a Mechanism Involving KatG and KatN.

Studies carried out in recent years have established that growth under conditions of reduced gravity enhances Salmonella enterica serovar Typhimurium virulence. To analyze the possibility that this microgravity-induced increase in pathogenicity could involve alterations in the ability of Salmonella to withstand oxidative stress, we have compared the resistance to hydrogen peroxide of various Salmonella enterica strains grown under conditions of low shear modeled microgravity (LSMMG) or normal gravity (NG). We have found that growth in LSMMG significantly enhances hydrogen peroxide resistance of all the strains analyzed.

Carbon monoxide binding to the heme group at the dimeric interface modulates structure and copper accessibility in the Cu,Zn superoxide dismutase from Haemophilus ducreyi: in silico and in vitro evidences.

X-ray absorption near-edge structure (XANES) spectroscopy and molecular dynamics (MD) simulations have been jointly applied to the study of the Cu,Zn superoxide dismutase from Haemophilus ducreyi (HdSOD) in interaction with the carbon monoxide molecule. The configurational flexibility of the Fe(II)-heme group, intercalated between the two subunits, has been sampled by MD simulations and included in the XANES data analysis without optimization in the structural parameter space. Our results provide an interpretation of the observed discrepancy in the Fe-heme distances as detected by extended X-ray absorption fine structure (EXAFS) spectroscopy and the classical XANES analysis, in which the structural parameters are optimized in a unique structure.

Fe-heme structure in Cu, Zn superoxide dismutase from Haemophilus ducreyi by X-ray absorption spectroscopy.

We have carried out an X-ray Absorption Spectroscopy (XAS) study of ferric, ferrous, CO- and NO-bound Haemophilus ducreyi Cu,ZnSOD (HdSOD) in solution to investigate the structural modifications induced by the binding of small gaseous ligands to heme in this enzyme. The combined analysis of EXAFS and XANES data has allowed us to characterize the local structure around the Fe-heme with 0.02A accuracy, revealing a heterogeneity in the distances between iron and the two histidine ligands which was not evident in the X-ray crystal structure.

Structural basis of heme binding in the Cu,Zn superoxide dismutase from Haemophilus ducreyi.

The Cu,Zn superoxide dismutase from Haemophilus ducreyi is characterized by the unique ability to bind heme at its dimer interface. Here we report the high-resolution crystal structures of this protein in the heme-loaded (holo) and heme-free (apo) forms. Heme is asymmetrically bound between the two enzyme subunits, where heme iron is coordinated by two histidine residues, His64 and His 124, provided by the two subunits.

Copper and zinc binding properties of the N-terminal histidine-rich sequence of Haemophilus ducreyi Cu,Zn superoxide dismutase.

The Cu,Zn superoxide dismutase (Cu,ZnSOD) isolated from Haemophilus ducreyi possesses a His-rich N-terminal metal binding domain, which has been previously proposed to play a copper(II) chaperoning role. To analyze the metal binding ability and selectivity of the histidine-rich domain we have carried out thermodynamic and solution structural analysis of the copper(II) and zinc(II) complexes of a peptide corresponding to the first 11 amino acids of the enzyme (H(2)N-HGDHMHNHDTK-OH, L). This peptide has highly versatile metal binding ability and provides one and three high affinity binding sites for zinc(II) and copper(II), respectively.

Regulatory and structural differences in the Cu,Zn-superoxide dismutases of Salmonella enterica and their significance for virulence.

Many of the most virulent strains of Salmonella enterica produce two distinct Cu,Zn-superoxide dismutases (SodCI and SodCII). The bacteriophage-encoded SodCI enzyme makes the greater contribution to Salmonella virulence. We have performed a detailed comparison of the functional, structural, and regulatory properties of the Salmonella SodC enzymes.

Periplasmic Cu,Zn superoxide dismutase and cytoplasmic Dps concur in protecting Salmonella enterica serovar Typhimurium from extracellular reactive oxygen species.

Several bacteria possess periplasmic Cu,Zn superoxide dismutases which can confer protection from extracellular reactive oxygen species. Thus, deletion of the sodC1 gene reduces Salmonella enterica serovar Typhimurium ability to colonize the spleens of wild type mice, but enhances virulence in p47phox mutant mice. To look into the role of periplamic Cu,Zn superoxide dismutase and into possible additive effects of the ferritin-like Dps protein involved in hydrogen peroxide detoxification, we have analyzed bacterial survival in response to extracellular sources of superoxide and/or hydrogen peroxide.

A role for Haemophilus ducreyi Cu,ZnSOD in resistance to heme toxicity.

The Cu,Zn superoxide dismutase (Cu,ZnSOD) from Haemophilus ducreyi is the only enzyme of this class which binds a heme molecule at its dimer interface. To explore the role of the enzyme in this heme-obligate bacterium, a sodC mutant was created by insertional inactivation. No difference in growth rate was observed during heme limitation.

X-ray absorption investigation of a unique protein domain able to bind both copper(I) and copper(II) at adjacent sites of the N-terminus of Haemophilus ducreyi Cu,Zn superoxide dismutase.

The N-terminal metal binding extension of the Cu,Zn superoxide dismutase from Haemophilus ducreyi is constituted by a histidine-rich region followed by a methione-rich sequence which shows high similarity with protein motifs involved in the binding of Cu(I). X-ray absorption spectroscopy experiments selectively carried out with peptides corresponding to the two metal binding regions indicate that both sequences can bind either Cu(II) or Cu(I). However, competition experiments demonstrate that Cu(II) is preferred by histidine residues belonging to the first half of the motif, while the methionine-rich region preferentially binds Cu(I) via the interaction with three methionine sulfur atoms.

Distinctive functional features in prokaryotic and eukaryotic Cu,Zn superoxide dismutases.

Bacterial and eukaryotic Cu,Zn superoxide dismutases show remarkable differences in the active site region and in their quaternary structure organization. We report here a functional comparison between four Cu,Zn superoxide dismutases from Gram-negative bacteria and the eukaryotic bovine enzyme. Our data indicate that bacterial dimeric variants are characterized by catalytic rates higher than that of the bovine enzyme, probably due to the solvent accessibility of their active site.