Tag Archives: ATPase activity

Calix[4]arene C-90 and its analogs activate ATPase of the myometrium myosin subfragment-1

R. D. Labyntseva1, O. V. Bevza1, K. V. Lytvyn1, M. O. Borovyk1,
R. V. Rodik2, V. I. Kalchenko2, S. O. Kosterin1

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: labyntseva@biochem.kiev.ua;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: vik@ioch.kiev.ua

Numerous female reproductive abnormalities are consequences of disorders in uterus smooth muscle (myometrium) contractile function. In this work, we described activators of ATPase, which could be used for development of effective treatments for correcting this dysfunction. Myosin ATPase localized in the catalytic domain of myosin subfragment-1 transforms a chemical energy deposited in macroergic bonds of ATP into mechanical movement. It was shown that сalix[4]arene C-90 and its structural analogs functionalized at the upper rim of macrocycle with four or at least two N-phenylsulfonуltrifluoroacetamidine groups, are able to activate ATP hydrolysis catalyzed by myometrium myosin subfragment-1. It was shown with the method of computer modeling that N-phenylsulfonуltrifluoroacetamidine groups of calix[4]arene C-90 interact with responsible for binding, coordination and the hydrolysis of ATP amino acid residues of myosin subfragment-1. The results can be used for further research aimed at using calix[4]arene C-90 and its analogs as pharmacological compounds that can effectively normalize myometrium contractile hypofunction.

Effect of electromagnetic field of extremely low frequency on ATPase activity of actomyosin

Yu. V. Tseyslyer, O. V. Shelyuk, V. S. Martynyuk, N. E. Nuryschenko

ESC Institute of Biology, Taras Shevchenko Kyiv National University, Ukraine;
e-mail: yuliya.tseysler@gmail.com;

The Mg2+/Ca2+ and K+-ATPase actomyosin activity of rabbit skeletal muscle was evaluated by the Fiske-Subbarow method during a five-hour exposition of protein solutions in electromagnetic field of extremely low frequency of 8 Hz and 25 µT induction. The results of the study of the ATPase activity of actomyosin upon electromagnetic exposure have shown statistically significant changes that are characterized by a rather complex time dynamics. After 1, 2 and 4 hours of exposure of protein solutions the effect of ELF EMF exposure inhibits the ATPase activity compared to control samples, which are not exposed to the magnetic field. By the third and fifth hours of exposure to the electromagnetic field, there is a significant increase in the ATPase activity of actomyosin. It should be noted that a similar pattern of change in enzyme activity was universal, both for the environment by Mg2+ and Ca2+, and in the absence of these ions in the buffer. This can evidence for Ca2+-independent ways of the infuence of electromagnetic field (EMP) on biologic objects. In our opinion, the above effects are explained by EMP influence on the dynamic properties of actomyosin solutions, which are based on the processes of spontaneous dynamic formation of structure.

ATPase activity of rabbit skeletal muscles actomyosin complex under ultrasound effect

K. O. Medynska, N. Ye. Nurishchenko, L. I. Pelyukh, O. V. Shelyuk

Taras Shevchenko Kyiv National University;
Educational and Scientific Centre “Institute of Biology”, Ukraine;
e-mail: medinkat@ukr.net

Influence of continuous and impulsive ultrasound 0.05; 0.2; 0.4; 0.7 and 1.0 W/cm2 on ATPase activity of rabbit skeletal muscle actomyosin has been investigated in this work. It has been shown that most changes of Mg2+,Ca2+-ATPase activity are observed under 0.2 and 0.4 W/cm2 continuous ultrasound. K+-ATPase activity is inhibited by continuous ultrasound of all intensities studied. Impulsive 2 and 10 ms ultrasound did not change the Mg2+,Ca2+-ATPase activity. While K+-activity is reliably changed only under impulsive 0.7 and 1.0 W/cm2 ultrasound that can be explained by the thermal effect. It has been determined, when studying­ the reconstructed actomyosin with sound troponin complex, that troponin complex is the most ultrasound sensitive constituent of actomyosin.

Calix[4]arene C-99 inhibits myosin ATPase activity and changes the organization of contractile filaments of myometrium

R. D. Labyntseva1, A. A. Bevza1, A. О. Lul’ko1, S. О. Cherenok2,
V. I. Kalchenko2, S. О. Kosterin1

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: labyntseva@biochem.kiev.ua;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kiev

Calix[4]arenes are cup-like macrocyclic (polyphenolic) compounds, they are regarded as promising molecular “platforms” for the design of new physiologically active compounds. We have earlier found that сalix[4]arenе C-99 inhibits the ATPase activity of actomyosin and myosin subfragment-1 of pig uterus іn vitro. The aim of this study was to investigate the interaction of calix[4]arene C-99 with myosin from rat uterine myocytes. It was found that the ATPase activity of myosin prepared from pre-incubated with 100 mM of calix[4]arene C-99 myocytes was almost 50% lower than in control. Additionally, we have revealed the effect of calix[4]arene C-99 on the subcellular distribution of actin and myosin in uterus myocytes by the method of confocal microscopy. This effect can be caused by reorganization of the structure of the contractile smooth muscle cell proteins due to their interaction with calix[4]arene. The obtained results demonstrate the ability of calix[4]arene C-99 to penetrate into the uterus muscle cells and affect not only the myosin ATPase activity, but also the structure of the actin and myosin filaments in the myometrial cells. Demonstrated ability of calix[4]arene C-99 can be used for development of new pharmacological agents for efficient normalization of myometrial contractile hyperfunction.

Calixarene methylene bisphosphonic acids as promising effectors of biochemical processes

S. V. Komisarenko1, S. O. Kosterin1, E. V. Lugovskoy1, V. I. Kalchenko2

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: kinet@biochem.kiev.ua;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: vik@ioch.kiev.ua

This interdisciplinary study, performed with participation of research workers of Palladin Institute of Biochemistry and Institute of Organic Chemist­ry of NAS of Ukraine, is devoted to analysis of biochemical effects of some calixarene methylene bisphosphonic acids (cyclic phenol oligomers) on two well-known biological phenomenons – Mg2+-dependent ATP hydrolysis (myosin subfragment-1 of myometrium smooth muscle was used as an example) and fibrin polymerization.
Calix[4]arene С-97 (calix[4]arene methylene bisphosphonic acids) is a macrocyclic substance, which contains intramolecular highly ordered lipophilic cavity formed by four aromatic rings, one of which is functionalized at the upper rim with methylene bisphosphonic group. At concentration of 100 µM, this substance was shown to effectively inhibit ATPase activity of pig myometrium myosin subfragment-1 (inhibition coefficient І0.5 = 83 ± 7 µM). At the same time, this calix[4]arene causes significant (vs. control) increase of myosin subfragment-1 hydrodynamic diameter, which may indicate formation of an intermolecular complex between calixa­rene and myosin head. Computer simulation methods (docking and molecular dynamics with addition of grid technologies) enabled to elucidate the grounds of intermolecular interactions between calix[4]arene С-97 and myometrium myosin subfragment-1, that involve hydrophobic, electrostatic and π-π-stacking interactions, some of which are close to the ATPase active centre. In view of the ability of calixarenes to penetrate into the cell and their low toxicity, the results obtained may be used as a basis for further development of a new generation of supramolecular effectors (starting from the above mentioned substances, in particular calix[4]arene С-97) for regulation of smooth muscle contractile activity at the level of ATP dependent actin-myosin interaction.
Calix[4]arenes bearing two or four methylenebisphosphonic acid groups at the macrocyclic upper rim have been studied with respect to their effects on fibrin polymerization. The most potent inhibitor proved to be calix[4]arene tetrakis-methylene-bis-phosphonic acid (C-192), in which case the maximum rate of fibrin polymerization in the fibrinogen + thrombin reaction decreased by 50% at concentrations of 0.52·10-6 M (IC50). At this concentration, the molar ratio of the compound to fibrinogen was 1.7 : 1. For the case of desAB fibrin polymerization, the IC50 was 1.26·10-6 M at a molar ratio of C-192 to fibrin monomer of 4 : 1. Dipropoxycalix[4]-arene bis-methylene-bis-phosphonic acid (C-98) inhibited fibrin desAB polymerization with an IC50 = 1.31·10-4 M. We hypothesized that C-192 blocks fibrin formation by combining with polymerization site ‘A’ (Aa17–19), which ordinarily initiates protofibril formation in a ‘knob-hole’ manner. This suggestion was confirmed by an HPLC assay, which showed a host–guest inclusion complex of C-192 with the synthetic peptide Gly-Pro-Arg-Pro, an analogue of site ‘A’. Further confirmation that the inhibitor was acting at the initial step of the reaction was obtained by electron microscopy, with no evidence of protofibril formation being evident. Calixarene C-192 also doubled both the prothrombin time and the activated partial thromboplastin time in normal human blood plasma at concentrations of 7.13·10-5 and 1.10·10-5 M, respectively. These experiments demonstrate that C-192 is a specific inhibitor of fibrin polymerization and blood coagulation and can be used for the design of a new class of antithrombotic agents.

Effect of fullerene C(60) on ATPase activity and superprecipitation of skeletal muscle actomyosin

K. S. Andreichenko1, S. V. Prylutska1, K. O. Medynska1, K. I. Bogutska1,
N. E. Nurishchenko1, Yu. I. Prylutskyy1, U. Ritter2, P. Scharff2

Joint Ukrainian-German Center on Nanobiotechnology
1Taras Shevchenko National University of Kyiv, Ukraine;
e-mail: prylut@ukr.net;
2Technical University of Ilmenau, Institute of Chemistry
and Biotechnology, Germany

Creation of new biocompatible nanomaterials, which can exhibit the specific biological effects, is an important complex problem that requires the use of last accomplishments of biotechnology. The effect of pristine water-soluble fullerene C60 on ATPase activity and superprecipitation reaction of rabbit skeletal muscle natural actomyosin has been revealed, namely an increase of actomyosin superprecipitation and Мg2+, Са2+– and K+-ATPase activity by fullerene was investigated. We conclude that this finding offers a real possibility for the regulation of contraction-relaxation of skeletal muscle with fullerene C60.

Protective effect of tiacalix[4]arene-tetrasulphonate on heavy metal inhibition of myometrium myosin subfragment-1 ATP-hydrolase activity

R. D. Labyntsevа1, O. V. Bevza1, A. A. Bevza1, A. M. Lulko1,
S. Kharchenko2, V. I. Kalchenko2, S. O. Kosterin1

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
е-mail: labyntseva@biochem.kiev.ua;  kinet@biochem.kiev.ua;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv;
е-mail: vik@ioch.kiev.ua

Heavy metals have a negative effect on the contractility of uterine smooth muscles (myometrium), these effects can lead to various pathologies of a women reproductive system. To overcome these effects the methods for correcting the myometrium contractile activity are  to be developed. Catalyzed by myosin ATPase ATP hydrolysis is the most important reaction in the molecular mechanism of myo­metrium contraction. We have found an inhibitory effect of 0.03-0.3 mM Ni2+, Pb2+ and Cd2+ on enzymatic hydrolysis of ATP by myosin subfragment-1 obtained from swine uterine smooth muscles. We have demonstrated that 100 µM thiacalix[4]arene-tetrasulphonate (C-798) recovered to the control level of ATPase activity of myosin subfragment-1 in the presence of heavy metal cations. One of the most probable mechanisms of C-798 corrective activity is based on its ability to chelate heavy metals, thus cations Pb, Cd and Ni can be removed from the incubation medium. Computer simulation has demonstrated that the protective effect of C-798 may also be the result of weakening the interaction of heavy metal ions with amino acid residues of the myosin molecule near the active site of ATP hydrolase. The obtained results can be used for further research aimed at assessing the prospects of thiacalix[4]arene-tetrasulfonate as pharmacological compounds.

Actomyosin ATPase activity of skeletal muscles and the markers of tissue damage in the blood of rats under prolonged chronic alcoholization

Yu. V. Tseyslyer1, О. M. Podpalova2, N. Е. Nurishchenko1, V. S. Маrtyniuk1

1ESC Institute of Biology, Taras Shevchenko National University of Kyiv, Ukraine;
e-mail: yuliya.tseysler@gmail.com,
2Bogomolets National Medical University, Kyiv, Ukraine;
e-mail: olgapodpalova@gmail.com

The activity of creatine kinase and indices of lipid metabolism in the blood and also actomyosin ATPase activity of skeletal muscles of rats under chronic 8-month alcohol abuse were investigated. It is shown that actomyosin K+-ATPase activity of skele­tal muscles increases from two months of ethanol use, but actomyosin Mg2+-ATPase activity decreases during 6-8 months of alcoholization. From two months of ethanol use the creatine kinase activity, as an enzyme marker of muscle tissue damage, statistically significantly increases during all the period of the animals alcoholization. The level of total lipid increases after two months of alcohol consumption (in blood plasma by 30% and in erythrocyte mass by 65%). For longer periods of alcoholization (4-8 months) the level of lipids remains almost the same, whereas in erythrocyte mass it does not differ from control values. The level of diene conjugates in the blood plasma reduces and the amount of ketone derivatives of fatty acid residues increases that points to the inhibition of some components of the antioxidant system that control detoxification of hydroperoxides of fatty acids and also to activation of free radical damage of tissues. There were no significant changes of lipid peroxidation level in erythrocyte mass at any stage of alcoholization.