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BMC Microbiology Apr 2023Environmental contamination by phenol has been reported in both aquatic and atmospheric environments. This study aimed to separate and purify the peroxidase enzyme from...
Environmental contamination by phenol has been reported in both aquatic and atmospheric environments. This study aimed to separate and purify the peroxidase enzyme from bacteria that degrade phenol from wastewater sources. An enrichment culture of MSM was used to screen 25 bacterial isolates from different water samples for peroxidase production, six of the isolates exhibited high levels of peroxidase enzyme activity. Qualitative analysis of peroxidase revealed that isolate No. 4 had the highest halo zones (Poly-R478: 14.79 ± 0.78 mm, Azure B: 8.81 ± 0.61 mm). The promising isolate was identified as Bacillus aryabhattai B8W22 by 16S rRNA gene sequencing with accession number OP458197. As carbon and nitrogen sources, mannitol and sodium nitrate were utilized to achieve maximum peroxidase production. A 30-h incubation period was used with pH 6.0, 30 °C, mannitol, and sodium nitrate, respectively, for maximal production of peroxidase. Purified peroxidase enzyme showed 0.012 U/mg specific activity, and SDS-PAGE analysis indicated a molecular weight of 66 kDa. The purified enzyme exhibits maximum activity and thermal stability at pH values of 4.0 and 8.0, respectively, with maximum activity at 30 °C and complete thermal stability at 40 °C. In the purified enzyme, the Km value was 6.942 mg/ml and the Vmax value was 4.132 mol/ml/hr, respectively. The results demonstrated that Bacillus aryabhattai B8W22 has promising potential for degrading phenols from various phenol-polluted wastewater sources.
Topics: Phenol; Peroxidase; Wastewater; RNA, Ribosomal, 16S; Phenols; Peroxidases; Hydrogen-Ion Concentration
PubMed: 37120512
DOI: 10.1186/s12866-023-02850-9 -
Microbiology Spectrum Feb 2022Heme-containing peroxidases are widely distributed in the animal and plant kingdoms and play an important role in host defense by generating potent oxidants....
Heme-containing peroxidases are widely distributed in the animal and plant kingdoms and play an important role in host defense by generating potent oxidants. Myeloperoxidase (MPO), the prototype of heme-containing peroxidases, exists in neutrophils and monocytes. MPO has a broad spectrum of microbial killing. The difficulty of producing MPO at a large scale hinders its study and utilization. This study aimed to overexpress recombinant human MPO and characterize its microbicidal activities and . A human HEK293 cell line stably expressing recombinant MPO (rMPO) was established as a component of this study. rMPO was overexpressed and purified for studies on its biochemical and enzymatic properties, as well as its microbicidal activities. In this study, rMPO was secreted into culture medium as a monomer. rMPO revealed enzymatic activity similar to that of native MPO. rMPO, like native MPO, was capable of killing a broad spectrum of microorganisms, including Gram-negative and -positive bacteria and fungi, at low nM levels. Interestingly, rMPO could kill antibiotic-resistant bacteria, making it very useful for treatment of nosocomial infections and mixed infections. The administration of rMPO significantly reduced the morbidity and mortality of murine lung infections induced by Pseudomonas aeruginosa or methicillin-resistant Staphylococcus aureus. In animal safety tests, the administration of 100 nM rMPO via tail vein did not result in any sign of toxic effects. Taken together, the data suggest that rMPO purified from a stably expressing human cell line is a new class of antimicrobial agents with the ability to kill a broad spectrum of pathogens, including bacteria and fungi with or without drug resistance. Over the past 2 decades, more than 20 new infectious diseases have emerged. Unfortunately, novel antimicrobial therapeutics are discovered at much lower rates. Infections caused by resistant microorganisms often fail to respond to conventional treatment, resulting in prolonged illness, greater risk of death, and high health care costs. Currently, this is best seen with the lack of a cure for coronavirus disease 2019 (COVID-19). To combat such untreatable microorganisms, there is an urgent need to discover new classes of antimicrobial agents. Myeloperoxidase (MPO) plays an important role in host defense. The difficulty of producing MPO on a large scale hinders its study and utilization. We have produced recombinant MPO at a large scale and have characterized its antimicrobial activities. Most importantly, recombinant MPO significantly reduced the morbidity and mortality of murine pneumonia induced by Pseudomonas aeruginosa or methicillin-resistant Staphylococcus aureus. Our data suggest that recombinant MPO from human cells is a new class of antimicrobials with a broad spectrum of activity.
Topics: Acute Disease; Animals; Anti-Infective Agents; Candida albicans; Drug Resistance, Bacterial; Escherichia coli; Female; HEK293 Cells; Humans; Hydrogen Peroxide; Male; Methicillin-Resistant Staphylococcus aureus; Mice; Mice, Inbred C57BL; Peroxidase; Pneumonia, Bacterial; Pseudomonas Infections; Pseudomonas aeruginosa; Recombinant Proteins; Staphylococcal Infections; Staphylococcus aureus
PubMed: 35019674
DOI: 10.1128/spectrum.00522-21 -
Journal of the American Society of... Aug 2022
Topics: Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis; Antibodies, Antineutrophil Cytoplasmic; Glomerulonephritis; Humans; Peroxidase; T-Lymphocytes
PubMed: 35906086
DOI: 10.1681/ASN.2022060668 -
Molecules (Basel, Switzerland) Jun 2023The sensitive and accurate determination of glyphosate (Glyp) is urgently demanded because it is closely correlated with human health and environmental safety. In this...
The sensitive and accurate determination of glyphosate (Glyp) is urgently demanded because it is closely correlated with human health and environmental safety. In this work, we proposed a sensitive and convenient colorimetric assay by employing copper ion peroxidases for the detection of Glyp in the environment. Free Cu(II) ions displayed high peroxidase activity and can catalytically oxidize the colorless 3,3',5,5'-tetramethylbenzidine (TMB) into blue oxTMB, resulting in an obviously visible discoloration reaction. Once the Glyp is added, the ability of copper ions to mimic peroxidase can be largely suppressed because of the generation of Glyp-Cu chelate. The favorable selectivity and sensitivity were demonstrated in the colorimetric analysis of Glyp. Furthermore, this rapid and sensitive method was successfully applied in the accurate and reliable determination of glyphosate in the real sample, holding promising applications in pesticide determination in the environment.
Topics: Humans; Colorimetry; Peroxidase; Copper; Oxidoreductases; Peroxidases; Ions; Hydrogen Peroxide; Glyphosate
PubMed: 37375185
DOI: 10.3390/molecules28124630 -
International Journal of Molecular... Dec 2022Implantation of scaffolds causes a local inflammatory response whereby the early recruitment of neutrophils is of great importance not only for fighting the infection,...
Implantation of scaffolds causes a local inflammatory response whereby the early recruitment of neutrophils is of great importance not only for fighting the infection, but also for facilitating effective regeneration. We used luminol-dependent chemiluminescence, flow cytometry, ELISA, and confocal microscopy to assess the responses of neutrophils after the exposure to the scaffold-decellularized bovine pericardium (collagen type I) crosslinked with genipin (DBPG). We demonstrated that DBPG activated neutrophils in whole blood causing respiratory burst, myeloperoxidase (MPO) secretion, and formation of neutrophil extracellular trap-like structures (NETs). In addition, we studied platelets, another important player of the immediate immune host response. We found that platelets triggered redox-activation of isolated neutrophils by the pericardium scaffold, and likely participate in the NETs formation. Free radicals generated by neutrophils and hypochlorous acid produced by MPO are potent oxidizing agents which can oxidatively degrade biological structures. Understanding the mechanisms and consequences of redox activation of neutrophils by pericardium scaffolds is important for the development of new approaches to increase the efficiency of tissue regeneration.
Topics: Cattle; Animals; Neutrophils; Extracellular Traps; Peroxidase; Oxidation-Reduction; Respiratory Burst; Blood Platelets
PubMed: 36555108
DOI: 10.3390/ijms232415468 -
ACS Applied Bio Materials Feb 2024In biosensor development, silk fibroin is advantageous for providing transparent, flexible, chemically/mechanically stable, biocompatible, and sustainable substrates,...
In biosensor development, silk fibroin is advantageous for providing transparent, flexible, chemically/mechanically stable, biocompatible, and sustainable substrates, where the biorecognition element remains functional for long time periods. These properties are employed here in the production of point-of-care biosensors for resource-limited regions, which are able to display glucose levels without the need for external instrumentation. These biosensors are produced by photopatterning silk films doped with the enzymes glucose oxidase and peroxidase and photoelectrochromic molecules from the dithienylethene family acting as colorimetric mediators of the enzymatic reaction. The photopatterning results from the photoisomerization of dithienylethene molecules in the silk film from its initial uncolored opened form to its pink closed one. The photoisomerization is dose-dependent, and colored patterns with increasing color intensities are obtained by increasing either the irradiation time or the light intensity. In the presence of glucose, the enzymatic cascade reaction is activated, and peroxidase selectively returns closed dithienylethene molecules to their initial uncolored state. Color disappearance in the silk film is proportional to glucose concentration and used to distinguish between hypoglycemic (below 4 mM), normoglycemic (4-6 mM), and hyperglycemic levels (above 6 mM) by visual inspection. After the measurement, the biosensor can be regenerated by irradiation with UV light, enabling up to five measurement cycles. The coupling of peroxidase activity to other oxidoreductases opens the possibility to produce long-life reusable smart biosensors for other analytes such as lactate, cholesterol, or ethanol.
Topics: Silk; Colorimetry; Peroxidases; Biosensing Techniques; Peroxidase; Glucose
PubMed: 38270977
DOI: 10.1021/acsabm.3c00872 -
Plant Physiology Apr 2023Reactive oxygen species are produced in response to pathogens and pathogen-associated molecular patterns, as exemplified by the rapid extracellular oxidative burst...
Reactive oxygen species are produced in response to pathogens and pathogen-associated molecular patterns, as exemplified by the rapid extracellular oxidative burst dependent on the NADPH oxidase isoform RESPIRATORY BURST OXIDASE HOMOLOG D (RBOHD) in Arabidopsis (Arabidopsis thaliana). We used the H2O2 biosensor roGFP2-Orp1 and the glutathione redox state biosensor GRX1-roGFP2 targeted to various organelles to reveal unsuspected oxidative events during the pattern-triggered immune response to flagellin (flg22) and after inoculation with Pseudomonas syringae. roGFP2-Orp1 was oxidized in a biphasic manner 1 and 6 h after treatment, with a more intense and faster response in the cytosol compared to chloroplasts, mitochondria, and peroxisomes. Peroxisomal and cytosolic GRX1-roGFP2 were also oxidized in a biphasic manner. Interestingly, our results suggested that bacterial effectors partially suppress the second phase of roGFP2-Orp1 oxidation in the cytosol. Pharmacological and genetic analyses indicated that the pathogen-associated molecular pattern-induced cytosolic oxidation required the BRI1-ASSOCIATED RECEPTOR KINASE (BAK1) and BOTRYTIS-INDUCED KINASE 1 (BIK1) signaling components involved in the immune response but was largely independent of NADPH oxidases RBOHD and RESPIRATORY BURST OXIDASE HOMOLOG F (RBOHF) and apoplastic peroxidases peroxidase 33 (PRX33) and peroxidase 34 (PRX34). The initial apoplastic oxidative burst measured with luminol was followed by a second oxidation burst, both of which preceded the two waves of cytosolic oxidation. In contrast to the cytosolic oxidation, these bursts were RBOHD-dependent. Our results reveal complex oxidative sources and dynamics during the pattern-triggered immune response, including that cytosolic oxidation is largely independent of the preceding extracellular oxidation events.
Topics: Arabidopsis Proteins; Peroxidase; Hydrogen Peroxide; Arabidopsis; NADPH Oxidases; Peroxidases; Plant Immunity; Mitochondria; Oxidation-Reduction; Oxidative Stress; Reactive Oxygen Species; Protein Serine-Threonine Kinases
PubMed: 36582183
DOI: 10.1093/plphys/kiac603 -
Sensors (Basel, Switzerland) Sep 2021Hypochlorous acid (HOCl) generates from the reaction between hydrogen peroxide and chloride ions via myeloperoxidase (MPO)-mediated in vivo. As very important reactive... (Review)
Review
Hypochlorous acid (HOCl) generates from the reaction between hydrogen peroxide and chloride ions via myeloperoxidase (MPO)-mediated in vivo. As very important reactive oxygen species (ROS), hypochlorous acid (HOCl)/hypochlorite (OCl) play a crucial role in a variety of physiological and pathological processes. However, excessive or misplaced production of HOCl/OCl can cause variety of tissue damage and human diseases. Therefore, rapid, sensitive, and selective detection of OCl is very important. In recent years, the fluorescent probe method for detecting hypochlorous acid has been developed rapidly due to its simple operation, low toxicity, high sensitivity, and high selectivity. In this review, the progress of recently discovered fluorescent probes for the detection of hypochlorous acid was summarized with the aim to provide useful information for further design of better fluorescent probes.
Topics: Fluorescent Dyes; Humans; Hydrogen Peroxide; Hypochlorous Acid; Peroxidase
PubMed: 34640646
DOI: 10.3390/s21196326 -
Journal of Microbiology and... Jan 2021γ-Glutamylcysteine synthetase (Gcs1) and glutathione reductase (Glr1) activity maintains minimal levels of cellular methylglyoxal in . In glutathione-depleted , we...
γ-Glutamylcysteine synthetase (Gcs1) and glutathione reductase (Glr1) activity maintains minimal levels of cellular methylglyoxal in . In glutathione-depleted , we previously saw that NAD(H)-linked methylglyoxal oxidoreductase (Mgd1) and alcohol dehydrogenase (Adh1) are the most active methylglyoxal scavengers. With methylglyoxal accumulation, disruptants lacking or exhibit a poor redox state. However, there is little convincing evidence for a reciprocal relationship between methylglyoxal scavenger genes-disrupted mutants and changes in glutathione-(in)dependent redox regulation. Herein, we attempt to demonstrate a functional role for methylglyoxal scavengers, modeled on a triple disruptant (//), to link between antioxidative enzyme activities and their metabolites in glutathione-depleted conditions. Despite seeing elevated methylglyoxal in all of the disruptants, the result saw a decrease in pyruvate content in // which was not observed in double gene-disrupted strains such as / and /. Interestingly, // exhibited a significantly decrease in HO and superoxide which was also unobserved in / and /. The activities of the antioxidative enzymes erythroascorbate peroxidase and cytochrome c peroxidase were noticeably higher in // than in the other disruptants. Meanwhile, Glr1 activity severely diminished in //. Monitoring complementary gene transcripts between double gene-disrupted / and / supported the concept of an unbalanced redox state independent of the Glr1 activity for //. Our data demonstrate the reciprocal use of Eapx1 and Ccp1 in the absence of both methylglyoxal scavengers; that being pivotal for viability in non-filamentous budding yeast.
Topics: Alcohol Dehydrogenase; Candida albicans; Cytochrome-c Peroxidase; Enzyme Assays; Fungal Proteins; Genes, Fungal; Glutamate-Cysteine Ligase; Glutathione; Hydrogen Peroxide; Oxidation-Reduction; Oxidoreductases; Peroxidase; Peroxidases; Pyruvaldehyde; Saccharomyces cerevisiae Proteins; Superoxides
PubMed: 33203822
DOI: 10.4014/jmb.2010.10057 -
Advanced Science (Weinheim,... Aug 2021Nanomaterials having enzyme-like activities are recognized as potentially important self-therapeutic nanomedicines. Herein, a peroxidase-like artificial enzyme is...
Nanomaterials having enzyme-like activities are recognized as potentially important self-therapeutic nanomedicines. Herein, a peroxidase-like artificial enzyme is developed based on novel biodegradable boron oxynitride (BON) nanostructures for highly efficient and multi-mode breast cancer therapies. The BON nanozyme catalytically generates cytotoxic hydroxyl radicals, which induce apoptosis of 4T1 cancer cells and significantly reduce the cell viability by 82% in 48 h. In vivo experiment reveals a high potency of the BON nanozyme for breast tumor growth inhibitions by 97% after 14-day treatment compared with the control, which are 10 times or 1.3 times more effective than the inert or B-releasing boron nitride (BN) nanospheres, respectively. This work highlights the BON nanozyme and its functional integrations within the BN nanomedicine platform for high-potency breast cancer therapies.
Topics: Animals; Antineoplastic Agents; Biocompatible Materials; Boron Compounds; Breast Neoplasms; Cell Line, Tumor; Cell Survival; Disease Models, Animal; Female; Mice; Mice, Inbred BALB C; Nanomedicine; Nanostructures; Peroxidase
PubMed: 34189868
DOI: 10.1002/advs.202101184