Acetylcholinesterase affinities

The reaction of a chemically active substance with a group located in an enzyme s active site, such that the stoichiometry of active sites can be determined analytically. See Acetylcholinesterase Affinity Labeling Diiso-propylfluorophosphate... 

AFFINITY ABELING ACTIVE-SITE TITRATION ACETYLCHOLINESTERASE AFFINITY ABELING DIISOPROPYLFLUOROPHOSPHATE ACTIVE TRANSPORT ION PUMPS... 

A two-site immunometric assay of undecapeptide substance P (SP) has been developed. This assay is based on the use of two different antibodies specifically directed against the N- and C-terminal parts of the peptide (95). Affinity-purified polyclonal antibodies raised against the six amino-terminal residues of the molecule were used as capture antibodies. A monoclonal antibody directed against the carboxy terminal part of substance P (SP), covalently coupled to the enzyme acetylcholinesterase, was used as the tracer antibody. The assay is very sensitive, having a detection limit close to 3 pg/mL. The assay is fiiUy specific for SP because cross-reactivity coefficients between 0.01% were observed with other tachykinins, SP derivatives, and SP fragments. The assay can be used to measure the SP content of rat brain extracts. 

While these functions can be a carried out by a single transporter isoform (e.g., the serotonin transporter, SERT) they may be split into separate processes carried out by distinct transporter subtypes, or in the case of acetylcholine, by a degrading enzyme. Termination of cholinergic neurotransmission is due to acetylcholinesterase which hydrolyses the ester bond to release choline and acetic acid. Reuptake of choline into the nerve cell is afforded by a high affinity transporter (CHT of the SLC5 gene family). 

Diazinon exerts its toxic effects by binding to the neuronal enzyme acetylcholinesterase (AChE) for long periods after exposure. Diazinon, in turn, is converted to diazoxon, which has a higher affinity for AChE (and thus greater toxicity) than the parent compound. There is a latent period in... 

Extraordinary selectivity has been accomplished with the parathion-type of insecticide by incorporating Q or CH3 groups in the meta position of the aryl ring. These groups interact sterically with acetylcholinesterase (AChE), increasing the affinity for the insect enzyme and decreasing it with the mammalian enzyme. Selectivity also is enhanced by differences in the rates of microsomal oxidation of P=S to P=0 and in hydrolytic detoxication between insects and mammals. The compounds, fenitrothion, fenthion, ronnel, bromophos, iodofenfos, chlorpyrifos—methyl, and pirimiphos —methyl, and dicapthon are used widely in public health, household, and agricultural pest control. 

Earlier work in this field [28] indicated that acetylcholinesterase enzymes would be suitable biomolecules for the purpose of pesticide detection, however, it was found that the sensitivity of the method varied with the type and source of cholinesterase used. Therefore the initial thrust of this work was the development of a range of enzymes via selective mutations of the Drosophila melanogaster acetylcholinesterase Dm. AChE. For example mutations of the (Dm. AChE) were made by site-directed mutagenesis expressed within baculovirus [29]. The acetylcholinesterases were then purified by affinity chromatography [30]. Different strategies were used to obtain these mutants, namely (i) substitution of amino acids at positions found mutated in AChE from insects resistant to insecticide, (ii) mutations of amino acids at positions suggested by 3-D structural analysis of the active site,... 

S. Andreescu, D. Fournier and J.-L. Marty, Development of highly sensitive sensor based on bioengineered acetylcholinesterase immobilized by affinity method, Anal. Lett., 36 (2003) 1865-1885. 

The acetylcholine, in turn, is hydrolyzed by both acetylcholinesterase and plasma butyryl-cholinesterase. Choline is actively transported into nerve terminals (synaptosomes) by a high-affinity uptake mechanism. Furthermore, the availability of choline regulates the synthesis of acetylcholine (Figure 14.4). 

Neurotransmitters are removed by translocation into vesicles or destroyed in enzyme-catalysed reactions. Acetylcholine must be removed from the synaptic cleft to permit repolarization and relaxation. A high affinity acetylcholinesterase (AChE) (the true or specific AChE) catalyses the hydrolysis of acetylcholine to acetate and choline. A plasma AChE (pseudo-AChE or non-specific AChE) also hydrolyses acetylcholine. A variety of plant-derived substances inhibit AChE and there is considerable interest in AChE inhibitors as potential therapies for cognition enhancement and for Alzheimer s disease. Organophosphorous compounds alkylate an active site serine on AChE and the AChE inhibition by this mechanism is the basis for the use of such compounds as insecticides (and unfortunately also as chemical warfare agents). Other synthetics with insecticidal and medical applications carbamoylate and thus inactivate AChE (Table 6.4). 

Camps, R Cusack, B. Mallender, W. D. El Achab, R. Morral, J. Munoz-Torrero, D. Rosenberry, T. L. Huprine X is a novel high-affinity inhibitor of acetylcholinesterase that is of interest for treatment of Alzheimer s disease. Mol. Pharmacol, 2000, 57(2) 409 17. 

There are different types of cholinesterases in the human body, which differ in their location in tissues, substrate affinity, and physiological function. The principal ones are acetylcholinesterase (AchE, EC 3.1.1.7), found in the nervous system and also present in the outer membrane of red blood cells, and plasma cholinesterases (EC 3.1.1.8, ChE), which are a group of enzymes present in plasma, liver, cerebrospinal fluid, and glial cells. Under normal physiological conditions, AChE performs the breakdown of acetylcholine (ACh), which is the chemical mediator... 

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