[A comparative study of the cholinesterase and carboxylesterase sensitivities of mammals and arthropods to new phenyl and glycidyl esters of dialkyl thiophosphoric acid]. / Sravnitel'noe issledovanie chuvstvitel'nosti kholinesteraz i karboksilésteraz nekotorykh mlekopitaiushchikh i chlenistonogikh k novym fenilovym i glitsidilovym éfiram dialkiltiofosfornoi kisloty.

The antienzymic activities of 14 organophosphorous compounds, the derivatives of dialkyl thiophosphoric acid, towards the acetylcholinesterase (AChE), butyrylcholinesterase (BuChE) and carboxylesterase (CE) from the spring grain aphid and mammals were investigated. The dependence of inhibitory activity of the compounds on their alkyl radical length was shown to be different for the AchE from the aphid and man. Some less pronounced differences in this dependence were revealed between the BuChEs from the aphid and horse as well as between the CEs from the aphid, mouse and red spider mite. The data give evidence of a distinction in structure of the active surfaces of the enzymes from the aphid and mammals. Some peculiar properties of the aphid cholinesterases are discussed taking account of the results of the present and previous papers.

[A comparative study of the substrate and inhibitor specificity of the glutathione transferase from the spring grain aphid (Schizaphis gramina Rond.) and from rat liver]. / Sravnitel'noe issledovanie substratnoi i ingibitornoi spetsifichnosti glutationtransferazy zlakovoi tli i pecheni krysy.

Kinetic parameters of 9 substrates interaction with glutathione transferase (GST) from spring grain aphid and rat were studied. The most significant difference in Vmax values was noticed for 4-nitropyridine-N-oxide (6 times higher for aphid) and ethacrynic acid (7 times higher for rat). Km values were practically in all cases higher for aphid GST as compared to rat GST. New class of effectors of GST suggested by us, that is azimines (2 series), was used for the inhibitor analysis. GST interaction with these inhibitors was appreciated by three types of activity: nucleophilic replacement, thiolysis and N-deoxygenation. It has been shown that the degree of GST inhibition depended considerably both on the GST source and the substrate used. New high-effective inhibitors of GST were found among azimines and their higher specificity to rat GST as compared to aphid GST was demonstrated especially in thiolysis reaction.

[Interaction of the rat liver glutathione transferase system with 1-phthalimidoazimines]. / Vzaimodeistvie glutationtransferaznoi sistemy pecheni krysy s 1-ftalimidoaziminami.

The interaction of rat liver glutathione and glutathione transferase with several 1-phthalimidoazimines is studied. Differences between spontaneous and enzymatic reactions of azimines and reduced glutathione are shown. It is pointed out that the formation of glutathione-azimine complexes takes place in the reaction mixture.

[Properties of cholinesterase preparations from human erythroyctes]. / Nekotorye svoistva preparatov atsetilkholinésterazy iz éritrotsitov cheloveka.

A modified method is described for isolation of acetylcholinesterase from human erythrocytes using an additional step of gel filtration on Sephadex G-75. Preparations of acetylcholinesterase were liberated from thromaboplastic activity and their specific activity was increased due to removal of low molecular proteins and of the products of destruction of hemoglobin. Content of A and B isoantigens in the preparations obtained was rather low and content of hemoglobin, combined with other proteins in the form of oxyhemoglobin, did not exceed 12% of the total protein.

[Ephedrine, salsoline and cytisine derivatives as substrates and inhibitirs of cholinesterases]. / Proizvodnye Efedrina, sal'solina i tsitizina v kachestve substratov i ingibitorov khokinesteraz.

25 iodomethylates of acetic, propionic, butyric, isobutyric and valeric esters of N-(beta-hydroxyethyl)-derivatives of ephedrine (I) pseudo-ephedrine (II), salsoline (III), salsolidine (IV) and cytisine (V) are studied as substrates and inhibitors of acetylcholine esterase (EC 3.1.1.8) from human erythrocytes and butyrylcholine esterase (EC 3.1.1.8) from horse serum. Butyrylcholine esterase found to increase the hydrolysis rate of all the alkaloid esters studied with the increase of acyl radical either to valerates (for ephedrine and pseudo-ephedrine derivatives), or to butyrates (for the rest alkaloids) and then it did not considerably change under further elongation of carbon chain up to valerate. Isobutyrates were observed to be similar to propionates in their hydrolysis rates. Acetylcholine esterase hydrolyzed acetates with the highest rate, while butyrates of ephedrine and pseudoephedrine derivatives were hydrolyzed by the enzyme 2,5-3-fold as slow as acetates. The rate of choline esterase hydrolysis decreased in the row: ephedrine--salsoline--cytisine with the volumetric increase of the cationic group. The decrease was almost 10-fold for butyrylcholine esterase, while a transition from "poor" substrates to reversible inhibitors was observed for acetylcholine esterase (3 of 5 cytisine esters were reversible inhibitors of the enzyme). The data obtained are compared with literary data on other cyclic choline esterase substrates; they are discussed from the viewpoint of unproductive binding hypothesis and on the basis of the structure of active centres of acetyl- and butyrylcholine esterases.

[Effect of Triton X-100 on properties of acetylcholinesterase from human erythrocytes]. / Vliianie tritona X-100 na svoistva atsetilkholinesterazy iz eritrotsitov cheloveka.

The effect of Triton X-100 on catalytic properties of acetylcholinesterase from human erythrocytes under acetylcholine hydrolysis, on sensitivity of acetylcholinesterase to specific phosphoorganic inhibitors and eserine, and on the mobility and isoenyme spectrum under analytical electrophoresis in polyacrylamide gel is investigated. Triton X-100, independently on its concentration within 0.05-1.0%, slightly changes V and [S]opt values and increases Km value in 2-3 times. The inhibitory effect of Triton X-100 is mainly competitive, 0.5% Triton X-100 decreases bimolecular constant (kII) of the interaction of acetylcholinesterase with phosphoorganic inhibitor and eserine in 2.5-4 times. In the presence of phosphoorganic inhibitor, kII sharply decreased when 0.02% Triton X-100 was added, and then it did not change under the increase of Triton X-100 concentration up to 1.0%. On the basis of these data, an analytical method of estimating Triton X-100 content in protein solution is proposed. The introduction of 0.1% Triton X-100 into polyacrylamide gel results in considerable quantitative redistribution of acetylcholinesterase isoenzyme fractions and in the change of the mobility of one fraction under electrophoresis.

[Comparative characteristics of acetylcholinesterase preparations from human erythrocytes with varying degrees of purity]. / Sravnitelnaia kharakteristika preparatov atsetilkholinésterazy iz éritrotsitov cheloveka razlichnoi stepeni ochistki.

Preparations of human erythrocyte acetyl cholinesterase (HEACE) that differed in their specific activities (0.7 to 4.1 U/mg) and the final purification method were examined for protein and isoenzymic spectrum (by polyacrylamide gel disc electrophoresis), catalytic properties in the reaction of acetytriocholine hydrolysis, sensitivity to the specific organophosphorus inhibitor Gd-42, concentrations of active centres of HEACE, and activity of one catalytic centre. The preparations showed a stable spectrum of 11-12 proteins: HEACE had molecular heterogeneity and was electrophoretically separated into 3 fractions that differed in their aggregation level. Preparations with varying specific activity contained different HEACE quantities. Regardless of the purification degree, HEACE displayed similar activity of the catalytic centre and enzymic properties in reactions with the substrate and inhibitor.

[Estimation of the irreversible inhibition reaction rate constants and of the concentration of cholinesterase active sites under commensurable concentrations of enzyme and inhibitors].

Kinetic analysis of the interaction of butyrylcholinesterase and the phosphoorganic inhibitor GT-161 [(C2H2O)2P(O)SC2H4+N(CH3)2C6H5-1-] is carried out. Short time incubations of an enzyme and an inhibitor (1-3 sec), even under commensurable concentrations, were shown to enable the rate constants of irreversible enzyme inhibition to be calcualted by the formula for pseudomonomoleuclar reactions. A simple analytical method for the estimation of the enzyme active sites concentrations is proposed.

[Determination of the rate constants for the interaction of enzymes with substrates and inhibitors during a short incubation time]. / Opredelenie konstnat skorostei vzaimodeistviia fermentov s substratami i ingibitorami za korotkoe vremia inkubatsii

Kinetics of enzymatic reactions was studied using equipment which permitted to decrease the time of incubation of an enzyme with substrate or inhibitor up to two sec; principles of action, design and experimental procedure are described. A method of pipetting under pressure was employed to accelerate an addition of suitable reagents to the enzyme solution and for rapid stirring of the reaction mixture. Time of incubation was automatically measured with a precision of plus or minus 0.01 sec using an electronic stop-watch. Start and stop of the watch were made by means of electric gauges with platinum electrodes, placed into the channels through which the substrate or inhibitor solutions were supplied to the reaction mixture.