Sensitivity of Cholinesterases of Different Origin to Organophosphorus Inhibitors with S-Alkyl Radical of Different Length

An antienzyme potency of 18 organophosphorus inhibitors (OPI), S-alkyl derivatives of thiophosphoric and thiophosphonic acids, to acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) of the spring grain aphid and corresponding mammalian enzymes is studied. It is shown that dependence of the inhibitory potency on length of alkyl radicals differs in human and aphid AChE. Smaller differences were observed in the case of aphid and horse BuChE. The obtained data are compared to the data obtained at studying other series of OPI and cholinesterases of other animals. It is suggested that the revealed peculiarity of the inhibitory specificity of aphid AChE is due to the presence of serine instead of phenylalanine in the position 331 (the numeration according to AChE of the electric ray Torpedo californica).     

Inhibitor Analysis of Cholinesterases of Different Origin (Variation of Structure of Leaving Group of Organophosphorus Inhibitors)

There is presented an analytic review of literature data on inhibitory analysis of structure of cholinesterases of different animals (vertebrates, insects, and molluscs) performed by studying rate of their interaction with 64 organophosphorus compounds of 13 series with different structure of leaving group. The presented data are considered from the point of view of comparative biochemistry and in the light of current data on structure of the cholinesterase active center.__________Translated from Zhurnal Evolyutsionnoi Biokhimii i Fiziologii, Vol. 41, No. 3, 2005, pp. 201–216.Original Russian Text Copyright © 2005 by Moralev, Rozengart.     

Comparative Sensitivity of Cholinesterases of Different Origin to Some Irreversible Inhibitors

The analytical review is presented of literature data obtained with use of different substrates, on sensitivity of more than 90 preparations of cholinesterases from 65 different animals (vertebrate, insects, molluscs) to the most studied 6 irreversible inhibitors (eserine and 5 organophosphorus compounds). The considered data are discussed from the point of view of comparative biochemistry and in the light of current information about structure of active center of cholinesterases.     

Ammonium Compounds with Localized and Delocalized Charge as Reversible Inhibitors of Cholinesterases of Different Origin

Five derivatives of benzimidazole, compounds with delocalized charge in cationic group, are studied and turned out to be reversible inhibitors of hydrolysis of acetylthiocholine under action of acetylcholinesterase from human erythrocytes, butyrylcholinesterase from horse blood serum, and cholinesterases from brain of the brown frog Rana temporaria and from optical ganglion of the Pacific squid Todarodes pacificus. It was only for acetylcholinesterase from erythrocyte as well as (with propyonylthiocholine as substrate) from squid that sensitivity to the studied benzimidazole derivatives correlated with degree of localization of the charge in the cationic group; this confirms the current concepts of functioning of the enzyme active center. A comparative study of 9 ammonium inhibitors with localized cation in their molecules, including the complete sterical analogue of the benzimidazole derivatives, benzimidazolinium iodide, has revealed both quantitative and qualitative differences.     

Indophenol Chromogenic Substrates of Cholinesterases of Different Origin

An analysis of influence of indophenol substrate structure on rate of their enzymatic hydrolysis under action of cholinesterases (ChE) of different animals is carried out for the first time. Study of indophenylacetate (IPhA) and a group of isomeric dichloroderivatives as substrates of erythrocyte acetylcholinesterase, serum butyrylcholinesterase, and ChE from optical ganglia of the Pacific squid Todarodes pacificus allowed us to reveal a role of steric and inductive effects of the substrates molecule in enzymatic catalysis, as well as differences in substrate specificity of the studied ChE. This comparative enzymologic aspect of the work was evident to a greater degree at studying hydrolysis of choline (acetylcholine, acetylthiocholine) and indophenol (IPhA, 2,6-dichloroindophenylacetate, 2,6-dichloro-3´-methyl indophenylacetate) esters under action of mammalian blood ChEs, ChE from hemolymph of the gastropod mollusc Neptunea, and also of ChE from the nervous tissue of different species of Pacific squids and of the cabbage root fly. Differences in values of the kinetic parameters characterizing sorption and catalytic stages of the hydrolysis process are revealed. Comparison of substrate properties of choline and indophenol esters enabled us to compare enzymes in terms of hydrophobic-hydrophilic interactions.     

Guanidine Derivatives: Conformation, Capability for Chelation, Study as Reversible Inhibitors of Cholinesterases of Different Origin

Study of spatial structure of biologically active guanidine derivatives by the method of molecular mechanics has shown that in an anticoccidial drug, 1,3-bis ( p-chlorobenzylidenamino)guanidine (Cl-BAG) the most preferable are convolute conformations, in which the chlorine atoms that are distant in the valent chain are approached to each other at a distance of 3.7 . This indicates predisposition of the optimal conformations to form chelate complexes with ions of metals, which is confirmed by comparative spectrophotometric studies of the second derivative of differential UV-spectra of Cl-BAG in the presence and absence of calcium ions. Its derivative without chlorine (BAG) is unable to bind Ca²⁺ and has been shown to have no anticoccidial action, which associates the biological potency with the presence of calcium-binding ability of the compounds. The capability of Cl-BAG for chelation depends essentially on nature of the chelated metal ion. The antienzyme testing of inhibiting action of the guanidine derivatives toward cholinesterases of human erythrocytes, horse blood serum, mink brain and serum, optic ganglia of the Pacific squid Todarodes pacificus has revealed difference between the enzymes due to possibility of redistribution of the positive charge between the guanidinium fragment and amino groups and a change of the degree of charge delocalization.     

Comparative Study of Reversible Organofluorine Ammonium Inhibitors of Cholinesterases of Different Animals

A group of organofluorine ammonium compounds, trimethyltrifluoromethylammonium, diethylmethyltrifluoromethylammonium, hexa(difluoromethylene)-bis(trimethylammonium), their non-substituted analogs as well as bis-onium organosilicone, phenyliodonium, and triphenylphosphonium derivatives were tested as reversible inhibitors of acetylcholinesterase of human erythrocytes, butyrylcholinesterase of horse blood serum, cholinesterase of brain of the frog Rana temporaria and cholinesterases of optic ganglion of the Pacific squid Todarodes pacificus. By the method of molecular mechanics, differences were revealed in conformational mobility of interonium chain and in geometric parameters of the studied compounds. It was shown that introduction of fluorine atoms into the inhibitor molecule affected only their interaction with the Pacific squid cholinesterase. It was possible to separate effects of the onium atom nature and of the interonium chain structure in the inhibitor molecule on the anticholinesterase potency.     

Use of Bis-Alkaloid Derivatives of Dicarboxylic Acids on the Basis of Lupinine, Anabasine and Cytisine as Reversible Inhibitors of Cholinesterases of Different Origin

Comparative study was carried out on action of a group of bis-alkaloid derivatives (on the basis of lupinine, anabasine, and cytosine, and their iodomethylates) of dicarboxylic acids (succinic, glutaric, azelaic, sebacic) on activity of cholinesterase of optical ganglia of the Commander squid Berryteuthis magister from different zones of habitation in the northwest part of the Pacific Ocean as well as human erythrocyte acetylcholinesterase and horse serum butyrylcholinesterase. These compounds turned out to be potent reversible inhibitors with specificity of action with respect both of sensitivity of the studied enzymes to them and of the type of their inhibitory action. The studied inhibitors can be used as tools in biochemical taxonomy to determine populational structure of such a marketing species as the Commander squid Berryteuthis magister.     

Non-Productive Binding of Substrates as One of Aspects of Substrate Specificity of Cholinesterases of Different Origin

Taking into account the phenomenon of non-productive binding of substrate, kinetic parameters of hydrolysis of acetylcholine (ACh) and its 13 derivatives with different structures of ammonium group by cholinesterase (ChE) of human erythrocytes, ChE of horse blood serum, and ChE of optic ganglia of the Pacific squid Todarodes pacificus are determined. A dependence is revealed of values of parameters of their enzymatic hydrolysis and parameters of the non-productive binding on the substrate structure and ChE nature. Effects of salts, LiCl, NaCl, KCl, MgCl₂, CaCl₂ and BaCl₂, on various kinetic parameters, including parameters of the non-productive binding of substrate, of enzymatic hydrolysis of iodides of ACh and N-acetoxyethylene-N-ethylpiperidinium under action of horse blood serum ChE are studied. Addition of the salts to the reaction mixture produced different effects on values of the catalytic center activity (a _c) and the Michaelis constant (K _M), depending on the cation nature and the substrate structure. At the same time, values of the a _c/K _M ratio that characterize to a degree the substrate affinity to the enzyme are equal to each other for two substrates differing in structure, regardless of the presence and nature of the studied cations. Parameters of the non-productive binding of N-acetoxyethylene-N-ethylpiperidinium iodide also depended on the salt nature; however, in that case, a question arises as to the correctness of the comparative analysis, when at determinations of the parameters the non-productive binding of ACh is ignored.     

Different Evolutionary Strategies for the Origin of Caspase-1 Inhibitors

Caspase 1 (CASP-1) inhibitors share sequence similarity to CASP-1 itself and are all mapped to chr11q22.3. Here we show that these inhibitors are all products of a series of gene duplications that occurred at this locus after the divergence between human and mouse. Surprisingly, stop codons originated independently in all duplicated copies to generate CARD-only proteins with inhibitory activity. We discuss this evolutionary model in the context of both neo- and subfunctionalization.     

Enzyme Inhibitors of Marine Microbial Origin with Pharmaceutical Importance

Several enzyme inhibitors with various industrial uses were isolated from bacteria and actinomycetes living in the marine environment. These inhibitors are useful in medicine and agriculture. All the compounds, except the monoamine oxidase inhibitors, are novel, and their activities have been characterized.