scholarly journals Harnessing Nature’s Diversity: Discovering organophosphate bioscavenger characteristics among low molecular weight proteins

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Reed B. Jacob ◽  
Kenan C. Michaels ◽  
Cathy J. Anderson ◽  
James M. Fay ◽  
Nikolay V. Dokholyan
Keyword(s):  
Binding Constants ◽  
Synaptic Cleft ◽  
Acetylcholinesterase Activity ◽  
Organophosphate Poisoning ◽  
Serine Hydrolase ◽  
Agricultural Pesticides ◽  
Protein Universe ◽  
Hydrolysis Of ◽  
Low Molecular Weight Proteins ◽  
Computational Strategy

Abstract 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 (kcat/Km of (2.0 ± 1.3) × 10−1 M−1s−1) and paraoxon (kcat/Km of (4.6 ± 0.8) × 103 M−1s−1), 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.

Cholinesterases and the Resistance of the Mouse Diaphragm to the Effect of Tubocurarine

2005 ◽  
Vol 103 (4) ◽  
pp. 788-795 ◽  
Author(s):  
Tu Nguyen-Huu ◽  
Alexandre Dobbertin ◽  
Julien Barbier ◽  
Jasmina Minic ◽  
Eric Krejci ◽  
...  
Keyword(s):  
Sucrose Gradient ◽  
Synaptic Cleft ◽  
Acetylcholinesterase Activity ◽  
Neuromuscular Blocking ◽  
Blocking Effect ◽  
Neuromuscular Blocking Agents ◽  
Twitch Response ◽  
Competitive Mechanism ◽  
Rate Of Hydrolysis ◽  
Hydrolysis Of

Background The diaphragm is resistant to competitive neuromuscular blocking agents. Because of the competitive mechanism of action of tubocurarine, the rate of hydrolysis of acetylcholine at the neuromuscular junction may modulate its neuromuscular blocking effect. The authors compared the neuromuscular blocking effect of tubocurarine on isolated diaphragm and extensor digitorum longus (EDL) muscles and quantified the acetylcholinesterase activity in hetero-oligomers. Methods Adult Swiss-Webster and collagen Q-deficient (ColQ) mice were used. The blocking effect of tubocurarine on nerve-evoked muscle twitches was determined in isolated diaphragm and EDL muscles, after inhibition of acetylcholinesterase by fasciculin-1, butyrylcholinesterase by tetraisopropylpyro-phosphoramide, or both acetylcholinesterase and butyrylcholinesterase by neostigmine, and in acetylcholinesterase-deficient ColQ muscles. The different acetylcholinesterase oligomers extracted from diaphragm and EDL muscles were quantified in sucrose gradient. Results The EC50 for tubocurarine to decrease the nerve-evoked twitch response was four times higher in the diaphragm than in the EDL. The activity of the different acetylcholinesterase oligomers was lower in the diaphragm as compared with the EDL. Inhibition of acetylcholinesterase by antagonists resulted in an increased dose of tubocurarine but an unchanged resistance ratio between the diaphragm and the EDL. A similar diaphragmatic resistance was found in ColQ muscles. Conclusion The current study indicates that, despite differences in acetylcholinesterase activity between the diaphragm and EDL, the diaphragmatic resistance to tubocurarine cannot be explained by the different rate of acetylcholine hydrolysis in the synaptic cleft.

scholarly journals Potential Anti-Acetylcholinesterase Activity of Cassia timorensis DC.

Molecules ◽  
2020 ◽  
Vol 25 (19) ◽  
pp. 4545
Author(s):  
Nurul Amira Nurul Azman ◽  
Maram B. Alhawarri ◽  
Mira Syahfriena Amir Rawa ◽  
Roza Dianita ◽  
Amirah Mohd Gazzali ◽  
...  
Keyword(s):  
Hydrogen Bond ◽  
Acetylcholinesterase Activity ◽  
The Other ◽  
Hydroxyl Group ◽  
Anionic Site ◽  
Methanol Extracts ◽  
Plant Parts ◽  
Ache Inhibitory Activity ◽  
First Time ◽  
Hydrolysis Of

Seventeen methanol extracts from different plant parts of five different Cassia species, including C. timorensis, C. grandis, C. fistula, C. spectabilis, and C. alata were screened against acetylcholinesterase (AChE). C. timorensis extracts were found to exhibit the highest inhibition towards AChE whereby the leaf, stem, and flower methanol extracts showed 94–97% inhibition. As far as we are aware, C. timorensis is one of the least explored Cassia spp. for bioactivity. Further fractionation led to the identification of six compounds, isolated for the first time from C. timorensis: 3-methoxyquercetin (1), benzenepropanoic acid (2), 9,12,15-octadecatrienoic acid (3), β-sitosterol (4), stigmasterol (5), and 1-octadecanol (6). Compound 1 showed moderate inhibition towards AChE (IC50: 83.71 μM), while the other compounds exhibited poor to slightly moderate AChE inhibitory activity. Molecular docking revealed that the methoxy substitution of 1 formed a hydrogen bond with TYR121 at the peripheral anionic site (PAS) and the hydroxyl group at C5 formed a covalent hydrogen bond with ASP72. Additionally, the OH group at the C3′ position formed an interaction with the protein at the acyl pocket (PHE288). This possibly explains the activity of 1 in blocking the entry of acetylcholine (ACh, the neurotransmitter), thus impeding the hydrolysis of ACh.

scholarly journals Effect of Cationic/Anionic Mixed Micelles on Reaction Kinetics of Alkaline Hydrolysis of Crystal Violet

2019 ◽  
Vol 31 (3) ◽  
pp. 651-655
Author(s):  
Qidist Yilma ◽  
Dunkana Negussa ◽  
Y. Dominic Ravichandran
Keyword(s):  
Crystal Violet ◽  
Alkaline Hydrolysis ◽  
Mixed Micelles ◽  
Sodium Dodecyl ◽  
Binding Constants ◽  
Regression Parameters ◽  
Experimental Values ◽  
Kinetics Of ◽  
Hydrolysis Of ◽  
Good Agreement

Kinetics of alkaline hydrolysis of crystal violet, a triphenylmethane dye in the micellar environment of cetyltrimethylammonium bromide (CTAB), sodium dodecyl sulfonate (SDS) and binary mixtures of these surfactants was studied. The regression parameters, together with rate constants and binding constants were obtained by analyzing the rate surfactant profiles using cooperativity model. It was observed that the reaction was catalyzed by both surfactants. The catalytic factor increased by 10 times in SDS and 38 times in CTAB indicating that binding of crystal violet to the micellar surface is stronger in pure CTAB than SDS but the strength drastically reduced in the mixtures of the surfactants. Reduction of binding constant became more important as the mole fraction of CTAB was improved in the mixture. The kinetic data were investigated using Piszkiewicz model and Raghavan-Srinivasan model. The data obtained from the models were in good agreement with the experimental values.

KINETIC MODELS ON ACETYLCHOLINESTERASE MODULATION BY SELF-SUBSTRATE AND POLYAMINES: ESTIMATION OF INTERACTION PARAMETERS AND RATE CONSTANTS FOR FREE AND ACETYLATED STATES OF THE ENZYME

2002 ◽  
Vol 10 (02) ◽  
pp. 127-147 ◽  
Author(s):  
ANDRÉS VENTURINO ◽  
ROSA MARÍA BERGOC ◽  
ANA MARÍA PECHEN DE D'ANGELO ◽  
ENRIQUE ARTURO ROSENBAUM
Keyword(s):  
Substrate Inhibition ◽  
Binding Constants ◽  
Acetylcholinesterase Activity ◽  
Affinity Constant ◽  
Amino Groups ◽  
Activation Free Energy ◽  
Interaction Factor ◽  
Quaternary Complex ◽  
Peripheral Site ◽  
Free Interaction

Polyamines act as dual modulators on electric eel acetylcholinesterase, modifying both the apparent Km and Ki, depending on substrate levels. A kinetic model was developed to explain the results, based on two-step catalysis, a peripheral site for substrate inhibition apart from the catalytic site, and one binding site for polyamine. This model presented the best fittings to data, when compared with a simpler one considering one catalytic step. A fitting equation built up with sixteen independent parameters let us calculate the kinetic constants. In this way, we were able to solve the parameter identifiability problem arising from model uncertainty when only substrate was used in acetylcholinesterase kinetics. Besides, fitting parameters directly provide information about the binding constants of the different complexes, the modulatory strength of substrate and polyamines, and the effect on the standard activation free energy for acetylcholinesterase. Substrate inhibition operates mainly on the first catalytic step with an affinity constant of 5.2 mM-1, which is reduced to one third for the acetylated enzyme. The interaction factor between substrate binding at both sites is about 12. The modulatory strength of polyamines is spermine > spermidine > putrescine. This order is directly related to the number of amino groups in the molecule, and to the calculated free interaction energy. The effect of the number of amino groups on the binding energy is significantly increased in acetylated acetylcholinesterase. It is also inferred that the formation of a quaternary complex enzyme-substrate-substrate-polyamine would not be possible. Some relations between polyamine structure and acetylcholinesterase activity are suggested from estimated constants. Due to the distal amino group distances, it is possible for spermine and spermidine to span along the catalytic gorge of acetylcholinesterase, binding to the catalytic and peripheral sites in a way similar to bisquaternary ammonium inhibitors.

scholarly journals Dramatic Increase in Cerebral Blood Flow following Soman Intoxication If Signs of Symptoms Can Be Seen

2015 ◽  
Vol 2015 ◽  
pp. 1-9 ◽  
Author(s):  
Ann Göransson Nyberg ◽  
Gudrun E. Cassel
Keyword(s):  
Blood Flow ◽  
Cerebral Blood Flow ◽  
Regional Blood Flow ◽  
Acetylcholinesterase Activity ◽  
Organophosphate Poisoning ◽  
Flow Measurements ◽  
Anaesthetized Rats ◽  
Compound Exposure ◽  
Treat All ◽  
Blood Flow Measurements

Organophosphate poisoning is associated with adverse effects on the central nervous system such as seizure/convulsive activity and long term changes in neuronal networks. This study report an investigation designed to assess the consequences of Soman, a highly toxic organophosphorus compound, exposure on regional blood flow in the rat brain and peripheral organs. We performed repeated blood flow measurements in the same animal, using the microspheres technique, to characterize changes in regional blood flow at different times after Soman intoxication. In addition, the cardiopulmonary effects of Soman were followed during the intoxication. Administration of Soman (1 LD50; 90 µg/kg, s.c.) to anaesthetized rats produced a decrease in blood acetylcholinesterase activity in all animals tested. Although, only six out of ten rats showed signs of poisoning like a decrease in respiratory rate, the results show that only animals with significant signs of poisoning demonstrated an increase in cerebral blood flow. We conclude that it is of great importance to treat all data individually. An overall mean can easily be misinterpreted and conceal important effects. We also conclude that the increase in cerebral blood flow has an important role in the effect on respiration and that this effect is independent of the blood acetylcholinesterase activity.

Neuromuscular Junction Disorders

2015 ◽  
Author(s):  
Anthony A Amato ◽  
Mohammad Kian Salajegheh
Keyword(s):  
Neuromuscular Junction ◽  
Myasthenia Gravis ◽  
Motor Nerve ◽  
Neuromuscular Disorders ◽  
Synaptic Cleft ◽  
Organophosphate Poisoning ◽  
Electric Currents ◽  
Repetitive Nerve Stimulation ◽  
Congenital Myasthenic Syndromes ◽  
Myasthenic Syndromes

The three main components of the neuromuscular junction (NMJ) include the presynaptic region, the synaptic cleft, and the postsynaptic region. The NMJ acts as an interface between the motor nerve and muscle by converting the motor nerve electric currents into chemical signals and then back into electric currents in the muscle. This chapter reviews electrodiagnostic testing in NMJ disorders, including repetitive nerve stimulation and single-fiber electromyography. Myasthenia gravis, congenital myasthenic syndromes, Lambert-Eaton myasthenic syndrome, botulism, and organophosphate poisoning and other toxins are discussed, including epidemiology, etiology/genetics, pathogenesis, diagnosis, differential diagnosis, management, complications, and prognosis. Tables include an overview of neuromuscular disorders, drugs with adverse effects on the NMJ, common immunomodulatory agents used for treatment of myasthenia gravis, congenital myasthenic syndromes, and toxins and venoms. Figures illustrate the NMJ structure and function, structure of the presynaptic and postsynaptic regions, electrodiagnostic studies in NMJ disorders, and dysfunction of the NMJ in acetylcholine receptor myasthenia gravis. This chapter contains 5 highly rendered figures, 5 tables, 65 references, and 5 MCQs.

scholarly journals Characterization and engineering of a two-enzyme system for plastics depolymerization

2020 ◽  
Vol 117 (41) ◽  
pp. 25476-25485 ◽  
Author(s):  
Brandon C. Knott ◽  
Erika Erickson ◽  
Mark D. Allen ◽  
Japheth E. Gado ◽  
Rosie Graham ◽  
...  
Keyword(s):  
Enzyme System ◽  
Environmental Crisis ◽  
Chimeric Proteins ◽  
Serine Hydrolase ◽  
Core Domain ◽  
Global Environmental ◽  
Pet Film ◽  
Lid Domain ◽  
Hydrolysis Of ◽  
Synergistic Relationship

Plastics pollution represents a global environmental crisis. In response, microbes are evolving the capacity to utilize synthetic polymers as carbon and energy sources. Recently,Ideonella sakaiensiswas reported to secrete a two-enzyme system to deconstruct polyethylene terephthalate (PET) to its constituent monomers. Specifically, theI. sakaiensisPETase depolymerizes PET, liberating soluble products, including mono(2-hydroxyethyl) terephthalate (MHET), which is cleaved to terephthalic acid and ethylene glycol by MHETase. Here, we report a 1.6 Å resolution MHETase structure, illustrating that the MHETase core domain is similar to PETase, capped by a lid domain. Simulations of the catalytic itinerary predict that MHETase follows the canonical two-step serine hydrolase mechanism. Bioinformatics analysis suggests that MHETase evolved from ferulic acid esterases, and two homologous enzymes are shown to exhibit MHET turnover. Analysis of the two homologous enzymes and the MHETase S131G mutant demonstrates the importance of this residue for accommodation of MHET in the active site. We also demonstrate that the MHETase lid is crucial for hydrolysis of MHET and, furthermore, that MHETase does not turnover mono(2-hydroxyethyl)-furanoate or mono(2-hydroxyethyl)-isophthalate. A highly synergistic relationship between PETase and MHETase was observed for the conversion of amorphous PET film to monomers across all nonzero MHETase concentrations tested. Finally, we compare the performance of MHETase:PETase chimeric proteins of varying linker lengths, which all exhibit improved PET and MHET turnover relative to the free enzymes. Together, these results offer insights into the two-enzyme PET depolymerization system and will inform future efforts in the biological deconstruction and upcycling of mixed plastics.

Cell membrane enveloped polymeric nanosponge for detoxification of chlorpyrifos poison: In vitro and in vivo studies

2021 ◽  
pp. 096032712199320
Author(s):  
S Altaf ◽  
F Muhammad ◽  
B Aslam ◽  
MN Faisal
Keyword(s):  
Glycolic Acid ◽  
Rabbit Model ◽  
Acetylcholinesterase Activity ◽  
In Vivo Studies ◽  
Organophosphate Poisoning ◽  
Neurotoxic Effects ◽  
Life Threatening ◽  
Red Blood Cell Membranes

Organophosphates are highly toxic compounds as they are involved in irreversible inhibition of acetylcholinesterase, causing various neurotoxic effects via acetylcholine accumulation throughout the nervous system. Traditional treatments for organophosphate poisoning are not effective enough to overcome all the toxic effects. There is a need for alternate treatment of life threatening poisoning of organophosphates. For this purpose a biomimetic nanosponge of poly (lactic- co-glycolic acid) is prepared, characterized and analysed as an antidote for organophosphate poisoning. In this nanosponge red blood cell membranes are used for coating poly lactic co-glycolic acid nanoparticles. In vitro studies are conducted to investigate the retention of acetylcholinesterase activity on the prepared nanosponge as well as to assess the scavenging ability of prepared nanosponge for model organophosphate, chlorpyrifos. In vivo studies are conducted to evaluate the detoxification potential of nanosponge in rabbit model, poisoned with chlorpyrifos. Hepatotoxicity and renal toxicity of nanosponge/chlorpyrifos complex is also studied in survived rabbits and the data is analysed statistically.

scholarly journals Isolation of Two Distinct Activator Proteins For Lipoprotein Lipase from Ovine Plasma

1987 ◽  
Vol 40 (3) ◽  
pp. 235
Author(s):  
RK Tume ◽  
RF Thornton ◽  
OW Johnson
Keyword(s):  
Lipoprotein Lipase ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Significant Inhibition ◽  
Activator Proteins ◽  
Human Apolipoprotein ◽  
Apolipoprotein C ◽  
Maximum Activation ◽  
Hydrolysis Of ◽  
Low Molecular Weight Proteins

Two distinct activator proteins for lipoprotein lipase (LPL) have been isolated in approximately equal amounts from ovine plasma. These low molecular weight proteins were readily separated from each other on the basis of size and charge. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated proteins of Mr about 8000 and 5000, with pI in urea-containing gels of about 5�1 and 4�8 respectively. Each of the ovine activator proteins was as effective as human apolipoprotein C-II (apo C-II) in activating LPL, with I JLg/ml giving near to maximum activation, and in lowering the apparent Km of LPL for triolein substrate. As the ratio of activator to triolein increased from 0�16 to 5� 2 (JLg/mg) the apparent Km fell from about O' 5 to 0 �18 mM. Whereas human apo C-II and the two ovine activators were equally effective in stimulating the hydrolysis of triolein, differences were observed between the human and ovine activators when p-nitrophenylbutyrate was used as substrate. Unlike human apo C-II, which produced significant inhibition of p-nitrophenylbutyrate hydrolysis, the ovine activators were without effect. This suggests that the interaction between the ovine activators and LPL is different from that of human apo C-II.

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