Defining the interplay of carbon dioxide and the carbonic anhydrase CanB in regulating Mycobacterium tuberculosis PhoPR signaling and virulence
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ABSTRACT:
ORGANISM(S): Mycobacterium tuberculosis
PROVIDER: GSE200125 | GEO | 2025/02/04
REPOSITORIES: GEO
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Project description:Carbonic anhydrases (CAs) are enzymes that catalyze the hydration/dehydration of CO2/HCO3(-) with rates approaching diffusion-controlled limits (kcat/KM ? 10(8) M(-1) s(-1)). This family of enzymes has evolved disparate protein folds that all perform the same reaction at near catalytic perfection. Presented here is a structural study of a ?-CA (psCA3) expressed in Pseudomonas aeruginosa, in complex with CO2, using pressurized cryo-cooled crystallography. The structure has been refined to 1.6 Å resolution with R(cryst) and R(free) values of 17.3 and 19.9%, respectively, and is compared with the ?-CA, human CA isoform II (hCA II), the only other CA to have CO2 captured in its active site. Despite the lack of structural similarity between psCA3 and hCA II, the CO2 binding orientation relative to the zinc-bound solvent is identical. In addition, a second CO2 binding site was located at the dimer interface of psCA3. Interestingly, all ?-CAs function as dimers or higher-order oligomeric states, and the CO2 bound at the interface may contribute to the allosteric nature of this family of enzymes or may be a convenient alternative binding site as this pocket has been previously shown to be a promiscuous site for a variety of ligands, including bicarbonate, sulfate, and phosphate ions.
| S-EPMC4745652 | biostudies-literature
Project description:The purpose of this study was to examine how ethoxzolamide modulates gene M. tuberculosis gene expression at acidic pH. We observed that ethoxzolamide downregulates genes of the PhoPR regulon.
2015-05-20 | GSE63917 | GEO
Project description:The purpose of this study was to examine how ethoxzolamide modulates gene M. tuberculosis gene expression at acidic pH. We observed that ethoxzolamide downregulates genes of the PhoPR regulon. Mtb strain CDC1551 was grown at 37C in T-25 vented, standing tissue culture flasks in 8 ml of buffered 7H9 medium seeded an initial OD of 0.2. Three conditions were examined with two biological replicates: 1) DMSO treated, pH 7.0, 2) DMSO treated, pH 5.7, 3) 40 µM ethoxzolamide treated, pH 5.7. The CDC1551(phoP::Tn) mutant was grown in a similar manner and treated with DMSO at pH 7.0 and 5.7. Following six days incubation, total bacterial RNA was extracted and analyzed as previously described.
2015-05-20 | E-GEOD-63917 | biostudies-arrayexpress
Project description:The visualization at near atomic resolution of transient substrates in the active site of enzymes is fundamental to fully understanding their mechanism of action. Here we show the application of using CO(2)-pressurized, cryo-cooled crystals to capture the first step of CO(2) hydration catalyzed by the zinc-metalloenzyme human carbonic anhydrase II, the binding of substrate CO(2), for both the holo and the apo (without zinc) enzyme to 1.1A resolution. Until now, the feasibility of such a study was thought to be technically too challenging because of the low solubility of CO(2) and the fast turnover to bicarbonate by the enzyme (Liang, J. Y., and Lipscomb, W. N. (1990) Proc. Natl. Acad. Sci. U. S. A. 87, 3675-3679). These structures provide insight into the long hypothesized binding of CO(2) in a hydrophobic pocket at the active site and demonstrate that the zinc does not play a critical role in the binding or orientation of CO(2). This method may also have a much broader implication for the study of other enzymes for which CO(2) is a substrate or product and for the capturing of transient substrates and revealing hydrophobic pockets in proteins.
| S-EPMC2576554 | biostudies-literature
Project description:Zinc tetraphenylporphyrin (Zn-TPP) solubilized by GroEL protein cage was prepared as a supramolecular mimic of carbonic anhydrase (CA) for CO2 capture. It is shown that the soluble Zn-TPP-GroEL complex can be formed easily by detergent dialysis. The Zn-TPP/GroEL binding ratio was found to increase with their dialysis ratio until reaching the maximum of about 30 porphyrins per protein cage. Moreover, the complex showed hydrase activity that catalyzes the CO2 hydration in HCO3- and H+. It is further seen that the catalytic activity of Zn-TPP-GroEL was about one-half of that of a bovine CA at 25 °C. On the other hand, as the temperature was increased to 60 °C close to an industrial CO2 absorption temperature, the natural enzyme lost function while Zn-TPP-GroEL exhibited better catalytic performance indicative of a higher thermal stability. Finally, we demonstrate that the GroEL-solubilized Zn-TPP is able to accelerate the precipitation of CO2 in the form of CaCO3 and has better long-term performance than the bovine CA. Thus a new type of nano-caged system mimicking natural CAs for potential applications in carbon capture has been established.
| S-EPMC7659338 | biostudies-literature
Project description:Carbon dioxide is vital to the chemistry of life processes including including metabolism, cellular homeostasis, and pathogenesis. CO2 forms carbamates on the neutral N-terminal a-amino- and lysine e-amino-groups that regulate the activities of ribulose 1,5-bisphosphate carboxylase/oxygenase and haemoglobin, however, very few protein other carbamates are known. Tools for the systematic identification of protein carbamylation sites have not been developed owing to the reversibility of carbamate formation, and in consequence carbamylation is typically overlooked. Here we demonstrate methods to identify protein carbamates using triethyloxonium ions to covalently trap CO2 on proteins for proteomic analysis. Our method delivers evidence to support the hypothesis that carbamylation is widespread in biology, and understanding its role should significantly advance our understanding of cellular CO2 interactions.
2018-10-23 | PXD007753 | Pride
Project description:Carbonic anhydrases are enzymes capable of transforming carbon dioxide into bicarbonate to maintain functionality of biological systems. Synthetic isolation and implementation of carbonic anhydrases into membrane have recently raised hopes for emerging and efficient strategies that could reduce greenhouse emission and the footprint of anthropogenic activities. However, implementation of such enzymes is currently challenged by the resulting membrane's wetting capability, overall membrane performance for gas sensing, adsorption and transformation, and by the low solubility of carbon dioxide in water, the required medium for enzyme functionality. We developed the next generation of enzyme-based interfaces capable to efficiently adsorb and reduce carbon dioxide at room temperature. For this, we integrated carbonic anhydrase with a hydrophilic, user-synthesized metal-organic framework; we showed how the framework's porosity and controlled morphology contribute to viable enzyme binding to create functional surfaces for the adsorption and reduction of carbon dioxide. Our analysis based on electron and atomic microscopy, infrared spectroscopy, and colorimetric assays demonstrated the functionality of such interfaces, while Brunauer-Emmett-Teller analysis and gas chromatography analysis allowed additional evaluation of the efficiency of carbon dioxide adsorption and reduction. Our study is expected to impact the design and development of active interfaces based on enzymes to be used as green approaches for carbon dioxide transformation and mitigation of global anthropogenic activities.
| S-EPMC8071118 | biostudies-literature
Project description:Colonoscopy is commonly used in screening for colorectal cancer. A refined technique of colonoscopy involving the use of water as the sole modality to aid colonoscope insertion, water exchange, has been described in recent research papers to decrease patient discomfort and pain, and to reduce the need for sedation during colonoscopy when compared with standard air insufflation. Carbon dioxide insufflation has been described to decrease patient discomfort after colonoscopy. No randomized trial has so far compared the use of water exchange to carbon dioxide insufflation. Our hypothesis is that water exchange inflicts less discomfort to patients undergoing colonoscopy than carbon dioxide insufflation. Patients undergoing screening colonoscopy in two centers in Norway, one center in Poland and one center in The Netherlands will be enrolled and randomized to examination of either of the two methods.
| 2128017 | ecrin-mdr-crc
Project description:Mycobacterium tuberculosis must sense and adapt to host environmental cues to establish and maintain an infection. The two-component regulatory system PhoPR plays a central role in sensing and responding to acidic pH within the macrophage and is required for M. tuberculosis intracellular replication and growth in vivo. Therefore, the isolation of compounds that inhibit PhoPR-dependent adaptation may identify new antivirulence therapies to treat tuberculosis. Here, we report that the carbonic anhydrase inhibitor ethoxzolamide inhibits the PhoPR regulon and reduces pathogen virulence. We show that treatment of M. tuberculosis with ethoxzolamide recapitulates phoPR mutant phenotypes, including downregulation of the core PhoPR regulon, altered accumulation of virulence-associated lipids, and inhibition of Esx-1 protein secretion. Quantitative single-cell imaging of a PhoPR-dependent fluorescent reporter strain demonstrates that ethoxzolamide inhibits PhoPR-regulated genes in infected macrophages and mouse lungs. Moreover, ethoxzolamide reduces M. tuberculosis growth in both macrophages and infected mice. Ethoxzolamide inhibits M. tuberculosis carbonic anhydrase activity, supporting a previously unrecognized link between carbonic anhydrase activity and PhoPR signaling. We propose that ethoxzolamide may be pursued as a new class of antivirulence therapy that functions by modulating expression of the PhoPR regulon and Esx-1-dependent virulence.
| S-EPMC4505220 | biostudies-literature
Project description:Carbon dioxide regulates cholesterol levels through SREBP2
| PRJNA804192 | ENA