[Correction of cardiac functions in rats with experimental myocardial ischemia by chronic administration of endothelin-converting enzyme inhibitor]. / Korrektsiia funktsional'nogo sostoianiia serdtsa krys s éksperimental'noi ishemiei miokada pri khronicheskom primenenii ingibitora éndotelin-prevrashchaiushchego fermenta.

Experiments on chronically instrumented Wistar rats demonstrated that 15 mm microsphere embolization of coronary arteries led to a significant decrease in the systemic (APsyst) and maximal left ventricular systolic pressures (LVSPmax) to 10.1 and 21.1%, respectively (p < 0.05). To evaluate the role of endothelin in this pathology, the inhibitor of endothelin-converting enzyme (ECE), PP-36, was used. PP-36 abolished hemodynamic changes caused by embolization after 4 days per os treatment (starting 2 days before surgical procedure). Dobutamine test revealed marked decrease of LVSPmax and +dP/dtmax responses in the embolized versus sham operated animals. PP-36 normalized ischemical heart response to beta-adrenergic stimulation. Maximal APsyst and LVSPmax increases were observed in embolized rats. PP-36 abolished this effect and led to parallel rising reaction to aminoguanidin in embolized (APsyst: +12.4 +/- 1.6 vs. +6.8 +/- 2.3 mmHg, p < 0.05) as well as in sham operated rats (APsyst: +8.5 +/- 1.1 vs. +5.6 +/- 0.7 mmHg, p < 0.05). Thus, the present research showed the possibility to correct ischemical heart disturbance by using a new ECE inhibitor, PP-36. One possible mechanism of this drugs action may include systemic or myocardial changes in NO contribution to the maintenance of normal arterial pressure.

[Study of phage T7 DNA-dependent RNA-polymerase using GTP analogs. Affinity modification and study of interaction with matrices using fluorescent markers]. / Issledovanie DNK-zavisimoi RNK-polimerazy faga T7 s pomoshch'iu analogov GTP. Affinaia modifikatsiia fluorestsentnymi metkami i issledovanie vzaimodeistviia s matritsei.

Interactions of the bacteriophage T7 DNA-dependent RNA polymerase with three GTP analogs have been studied. All of the three analogs tested contained substituted naphthalenesulphamide groups and were shown to be under appropriate conditions irreversible covalent inhibitors of the enzyme, the modified enzyme possessing fluorescent properties. One of these analogs contained the reactive 2-bromoethyl phosphonate group and was shown to cause the loss of the enzyme affinity for polynucleotide templates. The other two modifiers which contained the azide reactive group did not alter the enzyme-template affinity, the polynucleotide binding leading to a notable increase of the enzyme fluorescence intensity. The latter two modifiers are supposed to be convenient for fluorescent labelling of the active site of RNA polymerase for enzyme-template binding studies.

[The inhibiting effect of 8-Cl-adenosine-3',5'-cyclophosphate on the growth of melanoma B-16 in mice]. / Tormoziashchii éffekt 8-VCl-adenozin-3',5'-tsiklofosfata na rost melanomy B-16 u myshei.

A site-selective analogue of the cyclic adenosine monophosphate 8-chloro-adenosine-3',5'-cyclophosphate was studied for its effects on the growth of transplanted murine melanoma B-16. When the agent was given to the mice, a substantial effect on the growth of the tumor was produced by a number of factors, which included the route of administration, concentration of the agent, the time and duration of therapy. Intraperitoneal injections of the agent in a dose of 20 mg/kg/day which were made during three consecutive days, beginning from day 5 after tumor transplantation caused a 58% decrease in tumor growth as compared to the controls. An examination of tumour biopsy specimen revealed that after a course of the injections there was a significant suppression of the activity of cAMP-dependent protein kinase, type I, and a drastic increase in that of cAMP-dependent protein kinase, type II.

[Interaction of rabbit skeletal muscle glycogen synthase I with substrate analogs--oxo-UDP hydrazones]. / Vzaimodeistvie glikogensintazy I skeletnykh myshts krolika s analogamisubstrata--gidrazonami okso-UDP.

Inhibition of rabbit skeletal muscle glycogen synthase I was studied by using several synthetic substrate analogs: dansylhydrazone of oxo-UDP, 3-hydroxy-2-naphthoylhydrazone of oxo-UDP, salicyloylhydrazone of oxo-UDP, 1-oxyl-2,2,5,5-tetramethylpyrrolidine-3-carbonylhydrazone of oxo-UDP, N'-(dansyl)hydrazinocarbonylhydrazone of oxo-UDP and N'-(fluorenylidene-9)-hydrazinocarbonylhydrazone of oxo-UDP. All these compounds (with the exception of the nitroxyl-containing hydrazone) were characterized by a nonlinear dependence of the reverse reaction rate on the analog concentration in Dixon coordinates. The parabolic type of inhibition was due to the fact that the analogs tested except for the nitroxyl-containing hydrazone were able to interact both with the active center of the enzyme and with the FMN-binding site. The inhibition constants for oxo-UDP hydrazones were calculated for the both centers; their comparison revealed that the affinity of the analogs for the FMN-binding site increased with an increase in the radical hydrophobicity. These data suggest that the site with a high binding affinity for FMN is hydrophobic in nature. Apparently, isoalloxasine-like compounds display the highest affinity for this site.

[Effect of N-(6-aminohexyl)-5-chloronaphthalene-1-sulfamide (W-7) and its analogs on the activity of soluble guanylate cyclase and on human platelet aggregation]. / Vliianie N-(6-aminogeksil)-5-khlornaftalin-1-sul'famida (W-7) i ego analogov na aktivnost' rastvorimoi formy guanilattsiklazy i agregatsiiu trombotsitov cheloveka.

The effect of N-(omega-aminoalkyl) derivatives of naphthalene-1-sulfamide on the activity of soluble guanylate cyclase and on human platelet aggregation at the first (reversible) step of the guanylate cyclase reaction was studied. Low (approximately 10(-7)-10(-6) M) concentrations of the above compounds were shown to stimulate the guanylate cyclase activity; some derivatives caused simultaneous inhibition of platelet aggregation induced by ADP. Some fragments of the chemical structure of the molecules responsible for the enzyme activity regulation in the tested systems were identified. The naphthalene-1-sulfamide derivatives carrying 6-aminohexyl or 8-amino-octyl groups of the sulfamide substituent as well as chlorine atom at positions 4 or 5 of the naphthalene ring appeared to be the most potent activators of platelet guanylate cyclase and inhibitors of platelet aggregation at the reversible step of the enzymatic reaction.

[The study of nucleoside triphosphate-binding center of DNA-dependent RNA-polymerase of phage T7 using GTP analogs]. / Issledovanie nukleozidtrifosfatsviazyvaiushchego tsentra DNK- zavisimoi RNK-polimerazy faga T7 s pomoshch'iu analogov GTP.

The NTP binding site of bacteriophage T7 DNA-dependent RNA polymerase was studied using GTP analogs. For four analogs the irreversible inhibition was demonstrated. The kinetic parameters for competitive (Ki) and irreversible (KI and k3) inhibition were determined. One of the analogs, 5'[2-hydroxy(4-iodoacetamido)benzoyl]guanosine, was shown to inactivate the enzyme rapidly due to the modification of SH-groups. Some suggestions on the structure of the RNA polymerase active site have been made.

[Modulation of brain Ca2+-calmodulin-dependent phosphodiesterase activity by naphthalene sulfamides]. / Moduliatsiia fermentativnoi aktivnosti Ca2+-kalmodulin-zavisimoi fosfodiésterazy iz mozga soedineniiami naftalinsul'famidnogo riada.

Several W-7 analogs were synthesized. These compounds were shown to inhibit the calmodulin-activated phosphodiesterase. A comparative analysis of kinetic parameters of W-7 and its analogs was carried out. A hypothetical model of the effects of calmodulin inhibitors is proposed.

[Covalent modification of glycogen synthase I in rabbit skeletal muscle by 5'-fluorosulfonylbenzoyl derivatives of uridine]. / Kovalentnaia modifikatsiia glikogensintazy I skeletnykh myshts krolika 5'-ftorsul'fonilbenzoil'nymi proizvodnymi uridina.

Preincubation of rabbit skeletal muscle glycogen synthase I with 5'(p-fluorosulfonylbenzoyl)uridine (p-FSBU) or 5'-(m-fluorosulfonylbenzoyl)uridine (m-FSBU) results in a decrease of the enzymatic activity with time. UDP, the product of the glycogen synthase reaction, protects the enzyme against inactivation. It was shown that the covalent binding of the enzyme with each of its inhibitors is preceded by the formation of a reversible complex with Ki = 0.285 and 1.820 mM for p-FSBU and m-FSBU, respectively. This reaction is of pseudo-first order with respect to the inhibitors. The K2 values for p-FSBU and m-FSBU are nearly identical, i. e. 0.050 and 0.042 min-1, respectively, which suggests modification of the same functional group of the enzyme. The number and reactivity of the SH-groups of glycogen synthase I were determined, using 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB). The total number of SH-groups determined in the presence of 8 M urea is six as calculated per enzyme monomer. In the native enzyme DTNB titrates two SH-groups which according to their reactivity can be subdivided into 2 groups. The more reactive SH-group is localized in the active center of the enzyme. p-FSBU induces covalent blocking or screening of this SH-group during specific interaction with the active site of glycogen synthase I.

[Activation of adenyl cyclase from bovine brain caudate nucleus by synthetic analogs of guanyl nucleotides]. / Aktivatsiia adenilattsiklazy khvostatogo iadra mozga byka sinteticheskimi analogami guanilovykh nukleotidov.

In order to investigate the regulatory properties of dopamine-sensitive adenylate cyclase from bovine brain caudate nucleus some non-hydrolyzed guanyl nucleotides modified at the 8th position of the purine base and at phosphate site were synthesized. The activating effects of these compounds and that of GppCH2p on the membrane preparation of adenylate cyclase were studied. The activating effect of GppCH2p was decreased when the substituents were introduced at the 8th position of the purine base or when its terminal phosphate group was modified. The GDP analog ClCH2ppG-8-NH(CH2)6NHCOCH2Cl whose molecule contains simultaneously two substituents with alkylating chloromethyl groups produced an activating effect. This effect was abolished when the length of the substituent at the 8th position was reduced or one of the chloromethyl groups was lacking. The decrease of adenylate cyclase activation by ClCH2ppG-8-NH(CH2)NH(CH2)Cl in the presence of GDP is suggestive of a specific type of interaction of this compound with the enzyme. It is assumed that the components of the adenylate cyclase system may be covalently linked by this guanyl nucleotide.

[Irreversible inhibition of adenylate cyclase by oxo-ATP]. / Neobratimoe ingibirovanie adenilattsiklazy okso-ATP.

Adenylate cyclase from rabbit heart membranes is irreversibly inhibited by 2',3'-dialdehyde of ATP (oxo-ATP). The inhibiting effects is observed during membrane incubation with the inhibitor. Sodium borohydride increase the degree of the enzyme inactivation by oxo-ATP. The substrate protects the enzyme during incubation of the inhibitor with ATP. The data obtained suggest that the effect of oxo-ATP is localized in the enzyme active site and that the inhibitor blocks the amino group of the active site. The values of k2 and Ki for irreversible modification equal to 0.022 min-1 and 5.10(-4) M were determined.

[Adenylate cyclase from rabbit heart: substrate binding site]. / Adenilattsiklaza iz cerdtsa krolika: issledovanie substratsviazyvaiushchego uchastka.

The effects of 17 ATP analogs on the solubilized rabbit heart adenylate cyclase were studied. The triphosphate chain, position 8 of the adenine base and the ribose residue of the ATP molecule were modified. Despite the presence of the alkylating groups in two former types of the analogs tested, no covalent blocking of the active site of the enzyme was observed. Most of the compounds appeared to be competitive reversible inhibitors. The kinetic data confirmed the importance of the triphosphate chain for substrate binding in the active site of adenylate cyclase. (Formula: See Text) The inhibitors with different substituents in position 8 of the adenine base had a low affinity for the enzyme. The possible orientation of the triphosphate chain and the advantages of anti-conformation of the ATP molecule for their binding in the active site of adenylate cyclase are discussed.

[Some properties of the catalytic subunit of adenosine 3':5'-monophosphate-dependent protein kinase from pigeon breast muscle]. / Svoistva kataliticheskoi sub'edinitsy cAMP-zavisimoi proteinkinazy iz grudnoi myshtsy golubia.

A method for the preparation of a homogenous catalytic subunit of adenosine 3':5'-monophosphate-dependent protein kinase from pigeon breast muscle was developed. The molecular weight of the enzyme as determined by electrophoresis in the presence of sodium dodecyl sulfate was found to be 42000. The pH optimum of the catalytic subunit was around 8.0. The active site of the catalytic subunit was studied using some derivatives of ATP, containing different reactive groups in the triphosphate chain of the molecule. It may be assumed that the pH optimum of the enzyme inactivation by adenosine 5'-chloromethylpyrophosphonate and the protective effect of ATP suggest covalent binding of the imidazole ring in the enzyme active site. The kinetic mechanism of the protein kinase reaction was studied using the initial rate experiments and reaction product inhibition. The results obtained were consistent with a random Bi-Bi kinetic mechanism.

[A new method for synthesis of some nucleoside 5'-phosphonate esters]. / Novyi metod sinteza 5'-fosfonatnykh efirov nukleozidov.

A new method for synthesis of nucleoside-5'-phosphonates containing a reactive halogen atom in the phosphonate residue has been developed. In order to achieve selective inhibition of individual enzymes and to elucidate the structure of their active sites, the synthesis of 5-'chloromethyl-, 5'-(beta-bromoethyl)-and 5'-(N-chloroacetylaminomethyl)-phosphonate esters of adenosine and previously unknown analogous derivatives of uridine have been carried out. The method described may also be used to synthesize 5'-phosphonate esters of nucleosides, which contain radioactive labels.