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Molecules (Basel, Switzerland) May 2020We describe a patterned surface-enhanced Raman spectroscopy (SERS) substrate with the ability to pre-concentrate target molecules. A surface-adsorbed nanosphere...
Patterned Superhydrophobic SERS Substrates for Sample Pre-Concentration and Demonstration of Its Utility through Monitoring of Inhibitory Effects of Paraoxon and Carbaryl on AChE.
We describe a patterned surface-enhanced Raman spectroscopy (SERS) substrate with the ability to pre-concentrate target molecules. A surface-adsorbed nanosphere monolayer can serve two different functions. First, it can be made into a SERS platform when covered by silver. Alternatively, it can be fashioned into a superhydrophobic surface when coated with a hydrophobic molecular species such as decyltrimethoxy silane (DCTMS). Thus, if silver is patterned onto a latter type of substrate, a SERS spot surrounded by a superhydrophobic surface can be prepared. When an aqueous sample is placed on it and allowed to dry, target molecules in the sample become pre-concentrated. We demonstrate the utility of the patterned SERS substrate by evaluating the effects of inhibitors to acetylcholinesterase (AChE). AChE is a popular target for drugs and pesticides because it plays a critical role in nerve signal transduction. We monitored the enzymatic activity of AChE through the SERS spectrum of thiocholine (TC), the end product from acetylthiocholine (ATC). Inhibitory effects of paraoxon and carbaryl on AChE were evaluated from the TC peak intensity. We show that the patterned SERS substrate can reduce both the necessary volumes and concentrations of the enzyme and substrate by a few orders of magnitude in comparison to a non-patterned SERS substrate and the conventional colorimetric method.
Topics: Acetylcholinesterase; Carbaryl; Cholinesterase Inhibitors; Paraoxon; Spectrum Analysis, Raman
PubMed: 32397331
DOI: 10.3390/molecules25092223 -
Advances in Clinical and Experimental... 2013Paraoxonase-1 (PON1) is a HDL-attached extracellular esterase which is believed to contribute to the anti-atherogenic and anti-inflammatory properties of HDL. A decrease...
BACKGROUND
Paraoxonase-1 (PON1) is a HDL-attached extracellular esterase which is believed to contribute to the anti-atherogenic and anti-inflammatory properties of HDL. A decrease in PON1 is a risk factor for cardiovascular disease and has recently been found to be associated with juvenile obesity. The issue of a possible association between enzyme activity and/or its phenotype distribution and obesity-related metabolic abnormalities, inflammation, and oxidative stress has not been addressed yet.
OBJECTIVES
To evaluate PON1 activity and phenotype distribution with respect to obesity and obesity-related metabolic disorders, inflammation and oxidative stress in children and adolescents.
MATERIAL AND METHODS
PON1 arylesterase activity was measured spectrophotometrically in 156 children and adolescents (47 lean, 27 overweight and 82 obese). Enzyme phenotype was determined using dual substrate (phenyl acetate/paraoxon) method. PON1 activity and phenotype distribution were related to the presence of obesity, metabolic syndrome, insulin resistance, hyperinsulinemia, hypertriglyceridemia, high blood pressure, low HDL level, impaired fasting glucose and/or glucose tolerance as well as inflammatory and oxidative stress indices.
RESULTS
PON1 arylesterase activity decreased in general and central obesity, high blood pressure, and hyperinsulinemia conditions and correlated with BMI, CRP, adipocyte fatty acid-binding protein, superoxide dismutase, catalase, glutathione peroxidase, free thiols, and HOMA in a gender-dependent manner. PON1 decreases were independently associated with central obesity in girls, explaining 17% in PON1 variability, and with elevated CRP in boys, explaining 12% in its variability. PON1 phenotype was not associated with frequency of metabolic abnormalities.
CONCLUSIONS
PON1 decreases in central obesity, exacerbating obesity-related inflammation and oxidative stress. The enzyme associations are gender-dependent: obesity and oxidative stress affects PON1 in girls whereas inflammation in boys.
Topics: Adolescent; Aryldialkylphosphatase; Child; Enzyme Activation; Female; Humans; Inflammation; Male; Obesity; Overweight; Oxidative Stress; Risk Factors; Sex Characteristics; Sex Distribution; Substrate Specificity
PubMed: 23709379
DOI: No ID Found -
Sensors (Basel, Switzerland) Jan 2022The development of faster, sensitive and real-time methods for detecting organophosphate (OP) pesticides is of utmost priority in the in situ monitoring of these...
The development of faster, sensitive and real-time methods for detecting organophosphate (OP) pesticides is of utmost priority in the in situ monitoring of these widespread compounds. Research on enzyme-based biosensors is increasing, and a promising candidate as a bioreceptor is the thermostable enzyme esterase-2 from (EST2), with a lipase-like Ser-His-Asp catalytic triad with a high affinity for OPs. This study aimed to evaluate the applicability of Förster resonance energy transfer (FRET) as a sensitive and reliable method to quantify OPs at environmentally relevant concentrations. For this purpose, the previously developed IAEDANS-labelled EST2-S35C mutant was used, in which tryptophan and IAEDANS fluorophores are the donor and the acceptor, respectively. Fluorometric measurements showed linearity with increased EST2-S35C concentrations. No significant interference was observed in the FRET measurements due to changes in the pH of the medium or the addition of other organic components (glucose, ascorbic acid or yeast extract). Fluorescence quenching due to the presence of paraoxon was observed at concentrations as low as 2 nM, which are considered harmful for the ecosystem. These results pave the way for further experiments encompassing more complex matrices.
Topics: Biosensing Techniques; Ecosystem; Fluorescence Resonance Energy Transfer; Insecticides; Paraoxon; Pesticides
PubMed: 35062524
DOI: 10.3390/s22020561 -
Scientific Reports Nov 2016Organophosphate poisoning can occur from exposure to agricultural pesticides or chemical weapons. This exposure inhibits acetylcholinesterase resulting in increased...
Organophosphate poisoning can occur from exposure to agricultural pesticides or chemical weapons. This exposure inhibits acetylcholinesterase resulting in increased acetylcholine levels within the synaptic cleft causing loss of muscle control, seizures, and death. Mitigating the effects of organophosphates in our bodies is critical and yet an unsolved challenge. Here, we present a computational strategy that integrates structure mining and modeling approaches, using which we identify novel candidates capable of interacting with a serine hydrolase probe (with equilibrium binding constants ranging from 4 to 120 μM). One candidate Smu. 1393c catalyzes the hydrolysis of the organophosphate omethoate (k/K of (2.0 ± 1.3) × 10 Ms) and paraoxon (k/K of (4.6 ± 0.8) × 10 Ms), V- and G-agent analogs respectively. In addition, Smu. 1393c protects acetylcholinesterase activity from being inhibited by two organophosphate simulants. We demonstrate that the utilized approach is an efficient and highly-extendable framework for the development of prophylactic therapeutics against organophosphate poisoning and other important targets. Our findings further suggest currently unknown molecular evolutionary rules governing natural diversity of the protein universe, which make it capable of recognizing previously unseen ligands.
Topics: Data Mining; Databases, Protein; Hydrolysis; Organophosphates; Serine Endopeptidases
PubMed: 27845442
DOI: 10.1038/srep37175 -
International Journal of Molecular... Feb 2023Combined use of various antimicrobial peptides (AMPs) with enzymes that hydrolyze the signaling molecules of the resistance mechanism of various microorganisms, quorum...
Combined use of various antimicrobial peptides (AMPs) with enzymes that hydrolyze the signaling molecules of the resistance mechanism of various microorganisms, quorum sensing (QS), to obtain effective antimicrobials is one of the leading approaches in solving the antimicrobial resistance problem. Our study investigates the lactoferrin-derived AMPs, lactoferricin (Lfcin), lactoferampin and Lf(1-11), as potential partners for combination with enzymes hydrolyzing lactone-containing QS molecules, the hexahistidine-containing organophosphorus hydrolase (His-OPH) and penicillin acylase, to obtain effective antimicrobial agents with a scope of practical application. The possibility of the effective combination of selected AMPs and enzymes was first investigated in silico using molecular docking method. Based on the computationally obtained results, His-OPH/Lfcin combination was selected as the most suitable for further research. The study of physical-chemical characteristics of His-OPH/Lfcin combination revealed the stabilization of enzymatic activity. A notable increase in the catalytic efficiency of action of His-OPH in combination with Lfcin in the hydrolysis of paraoxon, -(3-oxo-dodecanoyl)-homoserine lactone and zearalenone used as substrates was established. Antimicrobial efficiency of His-OPH/Lfcin combination was determined against various microorganisms (bacteria and yeasts) and its improvement was observed as compared to AMP without enzyme. Thus, our findings demonstrate that His-OPH/Lfcin combination is a promising antimicrobial agent for practical application.
Topics: Quorum Sensing; Lactoferrin; Molecular Docking Simulation; Peptides; Anti-Infective Agents
PubMed: 36834977
DOI: 10.3390/ijms24043566 -
Journal of Microbiology and... Jan 2021Organophosphorus nerve agents (OPNAs), including both G- and V-type nerve agents such as sarin, soman, tabun and VX, are extremely neurotoxic organophosphorus compounds....
Organophosphorus nerve agents (OPNAs), including both G- and V-type nerve agents such as sarin, soman, tabun and VX, are extremely neurotoxic organophosphorus compounds. Catalytic bioscavengers capable of hydrolyzing OPNAs are under development because of the low protective effects and adverse side effects of chemical antidotes to OPNA poisoning. However, these bioscavengers have certain limitations for practical application, including low catalytic activity and narrow specificity. In this study, we generated a fusion-hybrid form of engineered recombinant human paraoxonase 1 (rePON1) and bacterial organophosphorus hydrolase (OPH), referred to as GV-hybrids, using a flexible linker to develop more promising catalytic bioscavengers against a broad range of OPNAs. These GV-hybrids were able to synergistically hydrolyze both G-type OPNA analogs (paraoxon: 1.7 ~ 193.7-fold, -nitrophenyl diphenyl phosphate (PNPDPP): 2.3 ~ 33.0-fold and diisopropyl fluorophosphates (DFP): 1.4 ~ 22.8-fold) and V-type OPNA analogs (demeton-Smethyl (DSM): 1.9 ~ 34.6-fold and malathion: 1.1 ~ 4.2-fold above) better than their individual enzyme forms. Among the GV-hybrid clones, the GV7 clone showed remarkable improvements in the catalytic activity toward both G-type OPNA analogs (/ (10 M min): 59.8 ± 0.06 (paraoxon), 5.2 ± 0.02 (PNPDPP) and 47.0 ± 6.0 (DFP)) and V-type OPNA analogs (/ (M min): 504.3 ± 48.5 (DSM) and 1324.0 ± 47.5 (malathion)). In conclusion, we developed GV-hybrid forms of rePON1 and bacterial OPH mutants as effective and suitable catalytic bioscavengers to hydrolyze a broad range of OPNA analogs.
Topics: Antidotes; Aryldialkylphosphatase; Catalysis; Genetic Engineering; Humans; Hydrolysis; Nerve Agents; Organophosphates; Organophosphorus Compounds; Phosphoric Triester Hydrolases; Recombinant Fusion Proteins; Substrate Specificity
PubMed: 33144547
DOI: 10.4014/jmb.2006.06044 -
Molecules (Basel, Switzerland) Jul 2017The albumin molecule, in contrast to many other plasma proteins, is not covered with a carbohydrate moiety and can bind and transport various molecules of endogenous and...
The albumin molecule, in contrast to many other plasma proteins, is not covered with a carbohydrate moiety and can bind and transport various molecules of endogenous and exogenous origin. The enzymatic activity of albumin, the existence of which many scientists perceive skeptically, is much less studied. In toxicology, understanding the mechanistic interactions of organophosphates with albumin is a special problem, and its solution could help in the development of new types of antidotes. In the present work, the history of the issue is briefly examined, then our in silico data on the interaction of human serum albumin with soman, as well as comparative in silico data of human and bovine serum albumin activities in relation to paraoxon, are presented. Information is given on the substrate specificity of albumin and we consider the possibility of its affiliation to certain classes in the nomenclature of enzymes.
Topics: Animals; Cattle; Enzyme Activation; Esterases; Humans; Hydrolysis; Ligands; Models, Molecular; Molecular Conformation; Organophosphates; Protein Binding; Serum Albumin; Substrate Specificity
PubMed: 28718803
DOI: 10.3390/molecules22071201 -
Journal of Thoracic Disease Aug 2018Knowledge on ontogenesis of thoracic surgery is essential not only for understanding present concepts and debates on surgery for tuberculosis, but it also contributes to... (Review)
Review
Knowledge on ontogenesis of thoracic surgery is essential not only for understanding present concepts and debates on surgery for tuberculosis, but it also contributes to the further developments in operative treatment of lung cancer. Both diseases have been the leading cause of death in their respective ages. History of tuberculosis follows the classic algorithm: diagnostic, casuistic and therapeutical stages. Villemin followed by Virchow, and, finally, Koch revealed the pathoanatomy and the cause of tuberculosis. The therapeutic phase of lung cancer has been reached without identified cause of the disease. Chest surgery, eradication of the macroscopic focus by physical interference with the involved tissue mass, in both diseases preceded medical treatment. Identification of phenotypes of lung cancer-if it is a single disease at all-does not contravene the concept: the tumor mass should been eliminated. However, causation is not an absolute sine qua non of an effective treatment, as the tuberculosis-lung cancer analogy also proves. Surgical approach of both diseases suffered from the same paraoxon: eradication without direct interference with the causative factor. While lung cancer seems to be controlled by an emerging array of new drugs, tuberculosis poses a new challenge, as multidrug resistant and extensively drug resistant Koch bacteria are emerging and fragile societies' immunity is weakening. Thoracic surgery has a significant share in the fight against tuberculosis, when drugs and/or society fail. Palliative and radical adjuvant surgery multiplies the chance of cure in those cases, where not much hope is left. The jury is still out in a series of questions, but it is obvious, that surgery is only an option and not a panacea where medicines and their providers fail. Deeper understanding of our past and present failures with tuberculosis and its surgery might contribute to new concepts in coping with lung cancer as well.
PubMed: 30345099
DOI: 10.21037/jtd.2018.04.131 -
Sensors (Basel, Switzerland) May 2022Developing an inexpensive, sensitive, and point-of-use biosensor for pesticide detection is becoming an important area in sensing. Such sensors can be used in food...
Developing an inexpensive, sensitive, and point-of-use biosensor for pesticide detection is becoming an important area in sensing. Such sensors can be used in food packaging, agricultural fields, and environmental monitoring of pesticides. The present investigation has developed a zinc oxide (ZnO)-based biosensor on porous, flexible substrates such as carbon paper and carbon cloth to detect organophosphates such as paraoxon (OP). Here, the influence of morphology and underlying substrate on biosensor performance was studied. The biosensors were fabricated by immobilizing the acetylcholinesterase (AChE) enzyme on ZnO, which is directly grown on the flexible substrates. The ZnO biosensors fabricated on the carbon cloth demonstrated good performance with the detection limit of OP in the range of 0.5 nM-5 µM, higher sensitivity, and greater stability.
Topics: Acetylcholinesterase; Biosensing Techniques; Carbon; Enzymes, Immobilized; Nanostructures; Paraoxon; Pesticides; Porosity; Zinc Oxide
PubMed: 35591210
DOI: 10.3390/s22093522 -
Tumour Biology : the Journal of the... Sep 2018Paraoxonase 1 plays an important role in protection from oxidative stress and also decomposes homocysteine thiolactone, the toxic metabolite of homocysteine. A limited...
Paraoxonase 1 plays an important role in protection from oxidative stress and also decomposes homocysteine thiolactone, the toxic metabolite of homocysteine. A limited number of reports evaluated the role of paraoxonase 1 in women affected by female genital tract neoplasms, including endometrial cancer. This study aimed to analyze the paraoxonase activity in the group of endometrial cancer patients (n = 48) who underwent primary surgery and to compare the data available with a well-matched control group (n = 30). Due to the role of paraoxonase 1 in the metabolism of homocysteine (Hcy) thiolactone, the amount of Hcy-thiolactone as well as total serum Hcy concentrations was also measured. Serum paraoxonase 1 activity toward synthetic substrates, paraoxon and phenyl acetate, in the study group was significantly lower compared to the control one. The mean paraoxonase 1 activity toward homocysteine thiolactone tended to be lower in the endometrial cancer group but this difference was not significant. There was no relationship between endometrial cancer and Q192R polymorphism of PON1 assessed by the dual substrate method. No differences in paraoxonase 1 activity between endometrial cancer subgroups according to clinico-pathological features were detected. Total serum homocysteine and protein-bound homocysteine thiolactone did not differ between control and cancer groups. In conclusion, reduced paraoxonase 1 activity suggests diminished important antioxidant mechanisms during the development of primary endometrial cancers in humans. PON1 Q192R polymorphism is not associated with the risk of endometrial cancer. Despite lower paraoxonase 1 activity, homocysteine concentration, and protein N-homocysteinylation in endometrial cancers do not differ from matched controls.
Topics: Aged; Aryldialkylphosphatase; Case-Control Studies; Endometrial Neoplasms; Female; Homocysteine; Humans; Lymphatic Metastasis; Middle Aged; Neoplasm Invasiveness; Prognosis; Protein Processing, Post-Translational
PubMed: 30178714
DOI: 10.1177/1010428318797869