The protein Pal (peptidoglycan-associated lipoprotein) is anchored in the outer membrane (OM) of Gram-negative bacteria and interacts with Tol proteins. Tol-Pal proteins form two complexes: the first ...is composed of three inner membrane Tol proteins (TolA, TolQ and TolR); the second consists of the TolB and Pal proteins linked to the cell's OM. These complexes interact with one another forming a multiprotein membrane-spanning system. It has recently been demonstrated that Pal is essential for bacterial survival and pathogenesis, although its role in virulence has not been clearly defined. This review summarizes the available data concerning the structure and function of Pal and its role in pathogenesis.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
The current therapies against gastric pathogen Helicobacter pylori are ineffective in over 20% of patients. Enzymes belonging to the purine salvage pathway are considered as novel drug targets in ...this pathogen. Therefore, the main aim of the current study was to determine the antibacterial activity of pyridoxal 5'-phosphate (PLP), an active form of vitamin B6, against reference and clinical strains of H. pylori. Using a broad set of microbiological, physicochemical (UV absorption, LC-MS, X-ray analysis) and in silico experiments, we were able to prove that PLP inhibits adenylosuccinate synthetase (AdSS) from H. pylori by the competition with GTP (IC50eq ∼30 nM). This behaviour was attributed to formation of a Schiff base with a lysine residue (a covalent bond with Lys322 in the GTP binding site of AdSS) and was potentiated by the presence of vitamin C. This antibacterial activity of PLP gives hope for its future use against H. pylori.The current therapies against gastric pathogen Helicobacter pylori are ineffective in over 20% of patients. Enzymes belonging to the purine salvage pathway are considered as novel drug targets in this pathogen. Therefore, the main aim of the current study was to determine the antibacterial activity of pyridoxal 5'-phosphate (PLP), an active form of vitamin B6, against reference and clinical strains of H. pylori. Using a broad set of microbiological, physicochemical (UV absorption, LC-MS, X-ray analysis) and in silico experiments, we were able to prove that PLP inhibits adenylosuccinate synthetase (AdSS) from H. pylori by the competition with GTP (IC50eq ∼30 nM). This behaviour was attributed to formation of a Schiff base with a lysine residue (a covalent bond with Lys322 in the GTP binding site of AdSS) and was potentiated by the presence of vitamin C. This antibacterial activity of PLP gives hope for its future use against H. pylori.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Posttranslational generation of disulfide bonds catalyzed by bacterial Dsb (disulfide bond) enzymes is essential for the oxidative folding of many proteins. Although we now have a good understanding ...of the Escherichia coli disulfide bond formation system, there are significant gaps in our knowledge concerning the Dsb systems of other bacteria, including Campylobacter jejuni, a food-borne, zoonotic pathogen. We attempted to gain a more complete understanding of the process by thorough analysis of C8J_1298 functioning in vitro and in vivo. C8J_1298 is a homodimeric thiol-oxidoreductase present in wild type (wt) cells, in both reduced and oxidized forms. The protein was previously described as a homolog of DsbC, and thus potentially should be active in rearrangement of disulfides. Indeed, biochemical studies with purified protein revealed that C8J_1298 shares many properties with EcDsbC. However, its activity in vivo is dependent on the genetic background, namely, the set of other Dsb proteins present in the periplasm that determine the redox conditions. In wt C. jejuni cells, C8J_1298 potentially works as a DsbG involved in the control of the cysteine sulfenylation level and protecting single cysteine residues from oxidation to sulfenic acid. A strain lacking only C8J_1298 is indistinguishable from the wild type strain by several assays recognized as the criteria to determine isomerization or oxidative Dsb pathways. Remarkably, in C. jejuni strain lacking DsbA1, the protein involved in generation of disulfides, C8J_1298 acts as an oxidase, similar to the homodimeric oxidoreductase of Helicobater pylori, HP0231. In E. coli, C8J_1298 acts as a bifunctional protein, also resembling HP0231. These findings are strongly supported by phylogenetic data. We also showed that CjDsbD (C8J_0565) is a C8J_1298 redox partner.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The genome of the strain
IBB3154 was sequenced, and transcriptome analysis was carried out at two different temperatures, allowing the determination of gene expression levels in response to ...environmental changes (temperature). Genes with higher expression at 42°C were identified. The use of a reporter gene (β- glucuronidase) did not confirm the transcriptomic results; it was found that the promoters of the genes
and
were active in the presence of bile salts. This opens up new opportunities for the overexpression of genes of other bacterial species in
cells in the intestinal environment.
Helicobacter pylori represents a global health threat with around 50% of the world population infected. Due to the increasing number of antibiotic-resistant strains, new strategies for eradication of ...H. pylori are needed. In this study, we suggest purine nucleoside phosphorylase (PNP) as a possible new drug target, by characterising its interactions with 2- and/or 6-substituted purines as well as the effect of these compounds on bacterial growth. Inhibition constants are in the micromolar range, the lowest being that of 6-benzylthio-2-chloropurine. This compound also inhibits H. pylori 26695 growth at the lowest concentration. X-ray structures of the complexes of PNP with the investigated compounds allowed the identification of interactions of inhibitors in the enzyme's base-binding site and the suggestion of structures that could bind to the enzyme more tightly. Our findings prove the potential of PNP inhibitors in the design of drugs against H. pylori.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
The bacterial proteins of the Dsb family catalyze the formation of disulfide bridges between cysteine residues that stabilize protein structures and ensure their proper functioning. Here, we report ...the detailed analysis of the Dsb pathway of
. The oxidizing Dsb system of this pathogen is unique because it consists of two monomeric DsbAs (DsbA1 and DsbA2) and one dimeric bifunctional protein (C8J_1298). Previously, we showed that DsbA1 and C8J_1298 are redundant. Here, we unraveled the interaction between the two monomeric DsbAs by in vitro and in vivo experiments and by solving their structures and found that both monomeric DsbAs are dispensable proteins. Their structures confirmed that they are homologs of EcDsbL. The slight differences seen in the surface charge of the proteins do not affect the interaction with their redox partner. Comparative proteomics showed that several respiratory proteins, as well as periplasmic transport proteins, are targets of the Dsb system. Some of these, both donors and electron acceptors, are essential elements of the
respiratory process under oxygen-limiting conditions in the host intestine. The data presented provide detailed information on the function of the
Dsb system, identifying it as a potential target for novel antibacterial molecules.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Lactic acid bacteria (LAB) are a diverse group of Gram-positive, nonsporulating, low G + C content bacteria. Many of them have been given generally regarded as safe status. Over the past two decades, ...intensive genetic and molecular research carried out on LAB, mainly Lactococcus lactis and some species of the Lactobacillus genus, has revealed new, potential biomedical LAB applications, including the use of LAB as adjuvants, immunostimulators, or therapeutic drug delivery systems, or as factories to produce therapeutic molecules. LAB enable immunization via the mucosal route, which increases effectiveness against pathogens that use the mucosa as the major route of entry into the human body. In this review, we concentrate on the encouraging application of Lactococcus and Lactobacillus genera for the development of live mucosal vaccines. First, we present the progress that has recently been made in the field of developing tools for LAB genetic manipulations, which has resulted in the successful expression of many bacterial, parasitic, and viral antigens in LAB strains. Next, we discuss the factors influencing the efficacy of the constructed vaccine prototypes that have been tested in various animal models. Apart from the research focused on an application of live LABs as carriers of foreign antigens, a lot of work has been recently done on the potential usage of nonliving, nonrecombinant L. lactis designated as Gram-positive enhancer matrix (GEM), as a delivery system for mucosal vaccination. The advantages and disadvantages of both strategies are also presented.
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CEKLJ, DOBA, EMUNI, GEOZS, IJS, IMTLJ, IZUM, KILJ, KISLJ, MFDPS, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UILJ, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
The Dsb protein family is responsible for introducing disulfide bonds into nascent proteins in prokaryotes, stabilizing the structure of many proteins. Helicobacter pylori HP0231 is a Dsb-like ...protein, shown to catalyze disulfide bond formation and to participate in redox homeostasis. Notably, many H. pylori virulence factors are stabilized by the formation of disulfide bonds. By employing H. pylori HP0231 deficient strains we analyzed the effect of lack of this bacterial protein on the functionality of virulence factors containing putative disulfide bonds. The lack of H. pylori HP0231 impaired CagA translocation into gastric epithelial cells and reduced VacA-induced cellular vacuolation. Moreover, H. pylori HP0231 deficient bacteria were not able to colonize the gastric mucosa of mice, probably due to compromised motility. Together, our data demonstrate an essential function for H. pylori HP0231 in gastric colonization and proper function of bacterial virulence factors related to gastric pathology.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Abstract Bacterial proteins of the Dsb (disulfide bond) system catalyze the formation of disulfide bridges, a post-translational modification of extra-cytoplasmic proteins, which leads to ...stabilization of their tertiary and quaternary structures and often influences their activity. DsbA – Escherichia coli monomeric oxidoreductase is the best studied protein involved in this process. Recent rapid advances in global analysis of bacteria have thrown light on the enormous diversity among bacterial Dsb systems. The set of Dsb proteins involved in the oxidative pathway, varies, depending on the microorganism. In this article we have focused on characterization of structural and phylogenetic groups of monomeric DsbAs. This review discuss their physicochemical features and interactions with redox partners as well as with substrate proteins. The last part of the review concentrates on dimeric oxidoreductases responsible for disulfide generation. Many virulence factors are the substrates of the Dsb proteins. Thus unraveling the machinery that introduces disulfide bonds and expanding knowledge about Dsb protein structures and their activities may facilitate the discovery of an effective anti-bacterial drugs. 1. Introduction. 2. Escherichia coli Dsb system. 2.1. Characteristic of the E. coli thiol oxidoreductase – DsbA. 2.2. Izomerization / reduction pathway proteins. 3. Classification of the monomeric DsbAs. 3.1. Physicochemical features of different classes of DsbAs. 4. DsbA interactions with redox partner and substrates. 4.1. DsbA interactions with redox partner. 4.2. DsbA interactions with substrates. 5. Dimeric Dsb proteins with oxidative activity. 6. Conclusions. 7. References
The recent, rapid increase in bacterial antimicrobial resistance has become a major public health concern. One approach to generate new classes of antibacterials is targeting virulence rather than ...the viability of bacteria. Proteins of the Dsb system, which play a key role in the virulence of many pathogenic microorganisms, represent potential new drug targets. The first part of the article presents current knowledge of how the Dsb system impacts function of various protein secretion systems that influence the virulence of many pathogenic bacteria. Next, the review describes methods used to study the structure, biochemistry, and microbiology of the Dsb proteins and shows how these experiments broaden our knowledge about their function. The lessons gained from basic research have led to a specific search for inhibitors blocking the Dsb networks.
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CEKLJ, DOBA, EMUNI, GEOZS, IJS, IMTLJ, IZUM, KILJ, KISLJ, MFDPS, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UILJ, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
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