[Scientific and practical activity of the Department of Muscle Biochemistry of the Palladin Institute of Biochemistry of NAS of Ukraine].

The article focuses on scientific and practical activity of the Department of Muscle Biochemistry of the Palladin Institute of Biochemistry of NAS of Ukraine in the context of its foundation and development. Main findings and practical achievements in the area of muscle biochemistry are summarized and discussed.

[Inventive activity of the Departments of Chemistry and Biochemistry of Enzymes, and Protein Structure and Function of the Palladin Institute of Biochemistry of NAS of Ukraine. Part III. Diagnostic test-systems for analysis of fibrinolysis blood system and novel approaches to thrombosis treatment].

This article continues analysis of scientific achievements of the Institute of Biochemistry in the study of hemostasis system. Two previous articles were focused on the studies of blood coagulation proteins and development of the immune-enzyme test-systems for evaluation of the risk of thrombosis upon various pathologies. This article highlights the research on the blood fibrinolysis system and new approaches to thrombosis treatment, which were developed (and are under development) in the Palladin Institute of Biochemistry of the NAS of Ukraine, in particular, in the Department of Chemistry and Biochemistry of Enzymes headed previously by Dr.Sci.(Biol.) S. O. Kudinov and now by Dr.Sci.(Biol.) T .V. Grinenko, and also in the Department of Protein Structure and Function headed by Dr.Biol.Sci. E. M. Makogonenko. The fundamental knowledge of protein molecule functions and mechanisms of regulation of blood coagulation and fibrinolysis opens up new opportunities to diagnose hemostasis disorders and control the effectiveness of the cardiovascular disease treatment and also contributes to development of new techniques for isolation of new proteins ­ promising therapeutic agents.

[Inventive activity of the Department of Molecular Immunology of the Palladin Institute of Biochemistry of NAS of Ukraine].

The article is devoted to the inventive activity of the Department of Molecular Immunology of the Palladin Institute of Biochemistry of NAS of Ukraine in the context of the history of its inception, development and in the context of scholarly and organizational activities of Sergii Vasyl'ovych Komisarenko. This autumn marks 50th anniversary since young Sergii Komisarenko (now ­ Academician of NAS and NAMS of Ukraine, Dr. Biol. Sci., Professor) has joined the Palladin Institute of Biochemistry, has completed all stages of the academic carrier from PhD student to Head of the Institute. He is the first in Ukraine who started the new branch of research ­ molecular immunology, created a strong scientific school, which earned worldwide acclaim and made significant contribution to finding solutions to current problems in human health sciences. S.V. Komisarenko was among those, who were first in the USSR to use immunoenzyme and flow cytofluometric assays, hybridoma technology for producing monoclonal antibodies and immunochemical assay of proteins, which became the basis for development of highly sensitive and highly specific immunodiagnostic systems, which are of high necessity in medicine, veterinary, development of immunotechnologies, environment monitoring, etc. Under his leadership the Department has made a series of important discoveries and developments including relating to antitumour immunotoxins, effects of low dose radiation on the immune system of Chernobyl liquidators, immunochemical structure of neurotoxin apamine, cytochrom c, fibrinogen and fibrin molecules at different stages of polymerization, diphtheria toxin and its receptor, tuberculosis causing micobacterium, roles of protease-activated receptors (PARs) and nicotinic acetylcholine receptors of lymphocytes, nature of polyreactive immunoglobulins (PRIGs), among other important scientific contributions. S.V. Komisarenko and his colleagues also hold numerous (more than 80) author's certificates and patents in Ukraine and USSR.

[Inventive activity of the Department of Metabolism Regulation of the Palladin Institute of Biochemistry of NAS of Ukraine].

The article is devoted to the inventive activity of the Department of Metabolism Regulation of the Palladin Institute of Biochemistry of NAS of Ukraine in the context of the history of its inception, development and the research activities of its founder, academician of NAS of Ukraine M. F. Guly as well as his students and followers. It briefly tells about practical achievements of M. F. Guly which were as significant, immense and diverse as his scientific accomplishments. The paper analyses in detail the practical results of scientific research of his students and followers aimed to solve practical problems of medicine, food-processing, agriculture, and which are essentially a continuation of the ideas and projects of M. F. Guly.

Stress fibres-a Ca2+ -independent store for annexins?

Annexins belong to a family of lipid-binding proteins that are implicated in membrane organization. Several members are capable of binding to actin and, in smooth muscle cells, annexin 6 is known to form a Ca(2+)-dependent, plasmalemmal complex with actin filaments. Annexins can also associate with F-actin containing stress fibres within cultured smooth muscle cells or fibroblasts in a Ca(2+)-independent manner. Depolymerization of stress-fibre systems with cytochalasin D leads to the translocation of actin-bound annexin 2 from the cytoplasm to the plasma membrane at high intracellular levels of Ca(2+). This type of Ca(2+)-dependent annexin mobility is observed only in cells of mesenchymal phenotype, which have a well-developed stress-fibre system; not in epithelial cells.

[ATP activity of myocardial actomyosin complex in patients with dilated cardiomyopathy]. / ATF-aznaia aktivnost' aktomiozinovogo kompleksa miokarda u bol'nykh s dilatatsionnoi kardiomiopatiei.

ATP activity of actomyosin in dilated cardiomyopathy (DCMP) was studied together with impact on the above activity of tropomyosin-troponin regulatory complexes recovered from the normal myocardium and cardiac muscle of a DCMP patient. Recordable in DCMP was a striking decline (1.5-fold) in ATP activity of actomyosin. But no significant difference was to be seen in sensitivity to Ca2+ ions of actomyosins obtained from the normal myocardium and cardiac muscle of the DCMP patient. The cardial tropomyosin-troponin regulatory complex from the DCMP patient's myocardium was shown to be endowed with somewhat more manifest activity compared to the analogous complex recovered from the normal myocardium.

Smooth muscle myosin light chain kinase, supramolecular organization, modulation of activity, and related conformational changes.

It has recently been suggested that activation of smooth muscle myosin light chain kinase (MLCK) can be modulated by formation of supramolecular structures (Sobieszek, A. 1991. Regulation of smooth muscle myosin light chain kinase. Allosteric effects and co-operative activation by CaM. J. Mol. Biol. 220:947-957). The present light scattering data demonstrate that the inactive (calmodulin-free) MLCK apoenzyme exists in solution as a mixture of oligomeric (2% by weight), dimeric (53%), and monomeric (45%) species at physiological ionic strength (160 mM salt). These long-living assemblies, the lifetime of which was measured by minutes, were in equilibrium with each other. The most likely form of the oligomer was a spiral-like hexamer, the dimensions of which fit very well the helical structure of self-assembled myosin filaments (Sobieszek, A. 1972. Cross-bridges on self-assembled smooth muscle myosin filaments. J. Mol. Biol. 70:741-744). After activation of the kinase by calmodulin (CaM) we could not detect any appreciable changes in the distribution of the kinase species either when the kinase was saturated with CaM or when its molar concentration exceeded that of CaM. Our fluorescent measurements suggest that the earlier observed inhibition of kinase at substoichiometric amounts of CaM (Sobieszek, A., A. Strobl, B. Ortner, and E. Babiychuk. 1993. Ca2+-calmodulin-dependent modification of smooth-muscle myosin light chain kinase leading to its co-operative activation by calmodulin. Biochem. J. 295:405-411) is associated with slow conformational change(s) of the activated (CaM-bound) kinase molecules. Such conformational rearrangements also took place with equimolar kinase to CaM; however, in this case there was no decrease in MLCK activity. The nature of these conformational changes, which are accompanied by reduction of the kinase for CaM affinity, is discussed.

Skeletal muscle myosin regulatory light chains conformation affects the papain cleavage of A1 light chains.

In the present study, the influence of magnesium-for-calcium exchange and phosphorylation of regulatory light chain (RLC) on accessibility of myosin and heavy meromyosin alkali light chains (A1) for papain digestion was investigated. The properties of native and papain treated myosin and heavy meromyosin were compared. Exchange of magnesium ions bound to RLCs for calcium ions accelerates the digestion of A1 in the presence of ATP in dephosphorylated myosin, heavy meromyosin, acto-myosin and the acto-heavy meromyosin complex. In the absence of ATP the exchange of magnesium ions bound to RLCs for calcium ions delays the digestion of A1 in the acto-myosin complex. Myosin and heavy meromyosin having shortened A1 by papain cleavage shows decreased K(+)-ATPase and increased actin binding ability in the presence and absence of ATP. The cooperation of RLC and A1 with heavy chains in the changes of structural organization of myosin head during muscle contraction is discussed.

[The localization of radioactive cesium in bovine muscle tissue]. / Izuchenie lokalizatsii radioaktivnogo tseziia v myshechnoi tkani krupnogo rogatogo skota.

The form in which radioactive caesium is present in muscular tissue has been investigated. It has been found that protein agents under study do not form stable complexes with radioactive caesium. It is suggested that caesium-137 is partially bound by muscular cell membranes upon meet storage.

Regulatory light chain influences alterations of myosin head induced by actin.

The effect of magnesium-for-calcium exchange and phosphorylation of regulatory light chain (LC2) on structural organization of rabbit skeletal myosin head was studied by limited tryptic digestion. In the presence of actin, exchange of magnesium bound to LC2 by calcium in dephosphorylated myosin accelerates the digestion of myosin and heavy meromyosin heavy chain and increases the accumulation of a 50 kDa fragment. This effect is significantly diminished in the case of phosphorylated myosin. Thus, both phosphorylation and cation exchange influences the effect of actin binding on the structural organization of myosin head.

Interaction of flavonoid compounds with contractile proteins of skeletal muscle.

Flavonoids (quercetin, rutin) influence ATPase activity and actomyosin superprecipitation. Low concentrations (below 20 mumol/l) of flavonoids were found to cause conformational changes in the myosin structure accompanied by an increase in ATPase activity. At higher concentrations an inhibitory action of flavonoids on both ATPase activity and actomyosin superprecipitation occurred. Conformational changes are likely to be due to flavonoids binding to regulatory site near the active centre of the myosin head. The effect of quercetin was stronger than that of rutin.

[Structural characteristics and aggregation of myosin heads in skeletal muscles]. / Strukturnye osobennosti i agregatsiia golovok miozina skeletnykh myshts.

Native conformational modifications of rabbit skeletal muscle myosin and its subfragment-1 (S-1) within the temperature range of 0-40 degrees C and irreversible unfolding of these proteins structure at temperatures 40-70 degrees C have been studied by the fluorescence and light scattering methods. The results obtained permit stating that myosin and its active subfragments form associates at the concentrations above 0.3 microM. Hydrophobic interactions between definite sites of S-1 are likely to be primarily responsible for the association. The complex profile of S-1 melting curve at high ionic strength indicates the existence of three structural domains in the heavy chain of the myosin head.

[Features of the preparation and estimation of homogeneity and native state of smooth muscle actin]. / Osobennosti polucheniia i otsenka gomogennosti i nativnosti aktina gladkikh myshts.

It is shown that smooth muscle actin preparations, produced with the help of techniques described in literature, have contaminations of tropomyosin and proteins with the molecular weight of 250 000, 80 000 and 18 000. The composition of contaminations depends on the method of preparation and conditions of dissociation of native actomyosin material. A method is proposed for actin separation from acetone-treated actomyosin followed by salting out with 50 mM MgCl2. Gel-electrophoresis in the presence of Na-dodecylsulphate has shown that the actin preparations are homogeneous and have the same cofactor activity for the myosin ATPase as actin from the rabbit skeletal muscle. It is supposed that in the cytoplasm of smooth muscle cells as well as in the non-muscle ones there is a considerable part of actin is the monomer form. It gets easily lost in the process of fibrillar actin separation. This may account for low actin output in spite of its relatively high contents in smooth muscles.