[Matrix activity of mRNP of different cell localization from plants infected by minus-genome curly potato dwarfness virus in protein-synthesizing system in vitro].

Using structures of mRNP of different cell localization isolated from plants infected by minus-genomecurly potato dwarfness virus (CPDV), it was established that synthesis of virus proteins in vitro is mainly realized on membrane-related and free polysomal mRNP capable to direct synthesis of proteins programmed in RNA in cell-free protein-synthesizing systems. The obtained results prove that synthesis of virus proteins is actively realized on membrane-related nonpolysomal mRNP - 23520 imp/min. An analogous distribution of matrix activity is observed also in the case when mRNA isolated from mRNP of different cell localization was introduced in protein-synthesizing system. The least matrix activity was observed in cytoplasmic nonpolysomal mRNPs which are low-active in the translation system in vitro - only 1890 imp/min. The function of free cytoplasmic nonpolysomal mRNP is apparently mainly reduced to the transport and reserve of genetic information in a cell.

[Effect of structural P-protein of potato curly dwarf virus on replicase activity of mRNP-complexes of infected plants].

All representatives of rhabdoviruses contain a nucleocapside phosphoprotein - P-protein which is an essential subunit of the viral RNA-dependent RNA polymerase complex. As a result of studying the effect of nucleocapside protein P(NS) on replicase activity of mRNP isolated from plants infected by potato curly dwarf virus in the system in vitro, it was established that nucleocapside P-protein stimulates considerably the replicase activity of membrane-bound polysomal m-RNP P-protein being available in concentration of 15 microg/ml in the replication system in vitro of membrane-bound polysomal mRNP, the replicase activity increased 11.7 times. This property of nucleocapside P-protein at the same concentration was displayed to a less extent with the presence of free polysomal mRNP, in the system in vitro. Thus the replicase activity mRNP-complexes in the replication system in vitro depends on the presence of nucleocapside viral P-protein in the system. Its concentration being increased or decreased, one can observe the change of the replicase activity.

[Inhibition of adenovirus reproduction in cell culture by specific antibodies].

The capacity of specific antibodies to inhibit the reproduction of homo- and heterologous adenoviruses in Hela cell added to culture medium after virus adsorption was studied. The inhibiting effect of polyclonal antivirus and monospecific antihexone antibodies to homo- and heterologous adenoviruses was shown. The effect was more expressed when using antibodies to homologous antibodies. The intensity of inhibition depended on antibodies concentration in the medium and infecting dose of the virus. Essential reduction of the quantity of infected cells and a decrease of the titer of adenovirus synthesized in the presence of homo- and heterologous antibodies was shown but adenovirus reproduction was not inhibited completely.

[Anti-adenovirus activity of a substance and medical form of ribamydil in cell culture].

The inhibiting effect of ribamydil on adenovirus reproduction was studied under the determination of the number of cells with virus- induced DNA-containing intranucleus inclusion bodies and hexone antigen, the synthesis of adenovirus proteins and the infection virus by t he investigation. EC50 of ribamydil substance is 4-8 microg/ml, but complete suppression of adenovirus genome expression was found when adding ribamydil after the virus adsorption, in concentrations of 125-500 microg/ml. The original effect of ribamydil on the expression of adenovirus genome was found under its effect in concentration of 31 microg/ml. Intranucleus virus-induced inclusion bodies of the early type only were found under these conditions. Synthesis of the structural virus polypeptides, including hexone polypeptide (II) and non-structural polypeptide 100K, taking part in hexone trimerization, proceed intensively but without formation of immunologically active hexone. The inhibiting effect of officinal form of ribamydil was less expressed as compared with the substance (EC50: 62 microg/ml). The work results prove that the therapeutic effect of ribamydil (ribavirin) under treatment of adenovirus infections may be achieved in case when it is used in a dose excluding the expression of the adenovirus genome.

[Research of modulation of CD95-mediated apoptosis in lymphoblastic MP-1 and BJAB cells infected by adenovirus and Epstein-Barr virus].

Model systems of infecting limphoblastic MP-1 and BJAB cells by Epstein-Barr virus, 5 serotype adenovirus and double infection are developed. A rather high level of accumulation of DNA of these viruses in the cells in dynamics at monoinfection and inhibition interference at multi-infection was shown by PCR method. The influence of virus infection on proliferative activity was studied. The stimulation of cells growth in the system BJAB + EBV was detected, and double infecting inhibited the process by 50%. The 25% difference in development of apoptosis process between cells infected by adenovirus and EBV was established when defining CD95-mediated apoptosis in infected MP-1 cells. The infecting of BJAB cells by viruses had a scarce effect on the processes of spontaneous apoptosis, but the data on CD95-mediated apoptosis at EBV infection testify to inhibition of this process both at a monoinfection, and at a double infection. The work was performed in the framework of the fundamental agreement of Ministry of Education and Science of Ukraine F7/366-2001, and grant INTAS N011-2382.

[Detection of antigen-antibody interaction of human adenovirus by the method of surface plasmon resonance].

A possibility to detect adenoviral protein--hexon, using specific antibodies by surface plasmon resonance (SPR) was demonstrated. The hexon of the human adenovirus 2 (Ad2) binds to antibodies immobilized on the sensor surface treated by KNCS and protein A Staphylococcus aureus. The specificity of antihexon antibodies was demonstrated by indirect method of fluorescent antibodies (MFA) and cellular variant of the immunoassay (cELISA).

[Functional activity of lymphoblastoid cells infected by human adenovirus type 2 and Epstein-Barr virus].

The paper deals with the influence of the adenovirus (Ad) and Epstein-Barr virus (EBV) on functional activity of lymphocytes, in particular, the production of alpha- and gamma-interferons, tumor necrosis factor (TNF) in conditions of mono- or double infection of B- and T-phenotype (CEM) lymphoblastoid cells. It is shown, that Ad, EBV or both viruses induce high enough levels of interferon on both lines of cells and in control epithelial cells. The lymphoblastoid cells infected by viruses deep ability to synthesize alpha- and gamma-interferons under the influence of the corresponding inducers (Newcastle disease virus and hemagglutinine). Nevertheless, the levels of their formation are not high. Rather high parameters of activity of the tumor necrosis factor (TNF) were revealed during a day in the initial B95-8 cells and superinfected Ad after the effect of LPS of E. coli. Their activity in CEM cells also did not depend on the infection type.

[Antigenic relationship between nucleocapsid proteins of phyto- and zoorhabdoviruses].

The methods of electrophoresis in PAAG and immunological method were used for comparative analysis of structural proteins of phytorhabdovirus of potato curly dwarf (PCDV) and zoorhabdoviruses-vesicular stomatitis virus (VSV) and fixed rabies Virus (RV). Molecular weight of viral proteins was determined by the method of polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The proteins with molecular weight 45-51 kD, are probably, the major component of the viral nucleocapsid. Nucleocapsid protein 45 kD RV virus was isolated by the method of preparative electrophoresis and then the monospecific serum was obtained. The Ouchterlony and immunoblotting method were used to show, that nucleocapsid proteins with molecular weights 51 and 45 kD both of phytorhabdovirus PCDV and zoorhabdoviruses VSV and RV are serologically related. The obtained data may be used in biotechnology as the basis for creation of a new class of diagnostic preparations with the purpose to detect RV virus using proteins of curly potato dwarf virus and may be also used in serological tests to reveal viruses of Rhabdoviridae family in various eukaryotic objects.

[Immunospecific detection of the potato virus X by the surface plasmon resonance].

Highly specific and proximal method of laboratory diagnostics of the viral disease, has been developed using the structural protein of the potato virus X, as a model, and monospecific antibodies to it. The immunospecific determination of the potato virus X was carried out by the surface plasmon resonance method using specific IgG-antigen complexes, immobilized on the sensor surface modified by rodanide and protein A of Staphylococcus aureus.

[Using the preparation "human immunoglobulin against herpes simplex virus type 1 for intramuscular injections" in the complex therapy of nervous system diseases]. / Zastosuvannia preparatu "imunohlobulin liudyny proty virusu herpesa zvychainoho 1-ho typu ridkyi dlia vnutrishn'om'iazovoho vvedenia" v kompleknomu likuvanni urazhen' nervovoï systemy.

The technology of obtaining of specific immunoglobulin for serotherapy of neuroinfection caused by virus herpes simplex 1 type was developed. The patients presented with the following diseases: arachnoencephalitis, encephalopolyradiculoneuritis, encephalomyelitis, encephalitis, arachnoiditis, polyneuropathy, encephalomyelopolyradiculoneuritis, meningoencephalitis. The study showed good tolerance and safety of the medicine, no adverse effects registered during the study. The assessed median score of the efficacy was 2.8 from 3. The obtained results suggest using the liquid form preparation for intramuscular injection "Immunoglobulin for treatment of neuroinfection caused by virus herpes simplex type 1". The Close corporation "Biofarma" located in Kyiv produces this medicine.

[Physico-chemical properties and functional peculiarities of mRNP of different subcellular location of Nicotiana rustica plants affected by potato curly dwarf virus]. / Fiziko-khimicheskie svoistva i funktsional'nye osobennosti mRNP razlichnoi subkletochnoi lokalizatsii rastenii Nicotiana rustica, porazhennykh virusom kurchavoi karlikovosti kartofelia.

mRNP of different subcellular location of N. rustica plants affected by PCDV contain virus-specific proteins. RNA-replicase, poly (A)-polymerase and protein kinase have been found in mRNP under study. The most expressed replicase, poly(A)-polymerase and protein kinase activity has been noticed in the systems containing membrane-bound non-polysomal mRNP.

[Enzyme activity of nucleocapsid proteins of the curly potato dwarf virus]. / Fermentativnaia aktivnost' nukleokapsidnykh belkov fitorabdovirusa kurchavoi karlikovosti kartofelia.

The individual nucleocapsid proteins of phytorhabdovirus of curly potato dwarf were isolated: N-protein (m. w. 56 kDa), NS-protein (m. w. 49 kDa), L-protein (m. w. 128 kDa). It was establish that L and NS proteins displayed enzyme activity in the replication, phosphorylation and adenylation systems in vitro. Major N protein (m. w. 56 kDa) did not show the enzyme activity in these systems. The revelation of RNA-polymerase, poly(A)-polymerase and proteinkinase activities of nucleocapsid proteins are characteristic of the viruses belonging to the Rhabdoviridae family.

[Investigation of interaction of immunoglobulins and detection of viral antigens in cell homogenates by the surface plasmon resonance method]. / Doslidzhennia vzaiemodiï imunohlobuliniv ta vyiavlennia virusnykh antyheniv u klitynnykh homohenatakh metodom poverkhnevoho plazmonnoho rezonansu.

Immobilization of immunoglobulins on the unmodified gold surface and the gold surface, modified by thiocyanate and protein A Staphylococcus aureus and their subsequent interaction with complementary antibodies were studied using the surface plasmon resonance method. Viral antigens were detected by surface plasmon resonance method in the cell homogenate of green alga Bracteococcus minor, which was artificially infected by tobacco mosaic virus. It was shown, that the process of interaction between virus and antiviral serum obey the Langmure model.

[Formation of research on viruses of humans, animals, and microorganisms at the Institute of Microbiology and Virology of the National Academy of Sciences of Ukraine and its present development]. / Stanovlennia doslidzhen' virusiv liudyny, tvaryn, mikroorhanizmiv v Instytuti mikrobiolohiï i virusolohiï NAN Ukraïny ta ïkh suchasnyi rozvytok.

The 40-year history of virology development at the D.K. Zabolotny Institute of Microbiology and Virology of the NAS of Ukraine and the basic success achieved in the study of human, insect viruses, phages of phytopathogenic bacteria have been analyzed.

[Method of orienting immobilized proteins on gold surface sensory modified with rhodanide]. / Sposob orientirovannoi immobilizatsii belkov na modifitsirovannykh rodanidom zolotykh poverkhnostiakh sensora.

The behavior of the biomolecules near the charged surface depending on the pH level of the solution was studied by the method of surface plasmon resonance. It was shown, that the preliminary modification of the surface by thiocyanat generating the effective negative charge at the surface allowed to immobilize protein molecule orientation depending on their charge state. Proteins keep their biological properties and form the oriented monolayer in the wide range of pH.

[Specific immunoglobulins as effective drugs for treatment of virus infections: present state and urgent tasks in Ukraine]. / Spetsificheskie immunoglobuliny kak éffektivnye preparaty dlia lecheniia virusnykh infekstii: sovremennoe sostoianie i nasushchnye zadachi dlia Ukrainy.

The thesis about importance of specific immunoglobulin preparations in treatment of infectious diseases of viral and microbial etiology, uncontrollable infections, caused by representatives of herpes viruses family in particular, is developed in the paper. The types of immunoglobulin preparations, their advantages, technology peculiarities has been considered; the experience of application, including that of the authors have been analyzed. The necessity and expedience of intensive development of this branch of immunobiotechnology in Ukraine is proved, especially in connection with a high level of infectious morbidity in this country, concrete recommendations are given.

[Orientation of proteins on the sensor surface and optimization of their immobilization by prior protection of the active center]. / Orientatsiia proteinov na poverkhnosti sensora i optimizatsiia ikh immobilizatsii putem predvaritel'noi zashchity aktivnogo tsentra.

When studying biospecific interactions with application of surface plasmon resonance, one of the main problems is reagent proper orientation to the sensor surface. Due to rather high chemical activity of molecular receptor sites, the interaction between these areas and surface may become predominant. Here we propose a technique for prevention of such orientation of bioreceptors using soybean trypsin inhibitor STI as an example. To obtain oriented STI immobilization on a modified gold surface its active site has been previously blocked through interaction with its specific partner trypsin. After conjugate immobilization on the sensor surface the components were separated using a glycine buffer (pH 2.2).

[Construction of the immunoenzyme test system "IFA-AtEBV-Strip" for detecting antibodies of the Epstein-Barr virus]. / Konstruiuvannia imunofermentnoï test-systemy "IFA-AtVEB-Strip" dlia vyiavlennia antytil proty virusu Epshteina-Barr.

The first home immunoenzyme test-system for finding antibodies to the Epstein-Barr virus (EBV) has been created. The test-system laboratory prototype was called "IFA-AtEBV-strip". Production technology has been developed for specific components of immunoenzyme test system and all the analysis stage conditions have been optimized: the antigen sorption on plates, its interaction with the studied serum, finding out of the formed immune complex. The test system "IFA-AtEBV-strip" sensitivity made up 96%, specificity--100%.

[The patterns in the molecular biology of the adenoviruses (the results of a 20-year study at the Institute of Microbiology and Virology of the National Academy of Sciences of Ukraine)]. / Nekotorye zakonomernosti molekuliarnoi biologii adenovirusov (itogi 20-letnego izlucheniia v Institute mikrobiologii i virusologii NAN Ukrainy).

Scientific data of the highest importance and priority concerning regularities of structural and functional organization of proteins of adenoviruses capsids and peculiarities of expression of the virus genome are as follows: New antigen determinants of hexon and adenovirus fiber have been discovered, their different nature (conformational or linear) and different orientation, depending on the spatial organization of proteins, have been proved; localization of some epitopes has been determined with the help of synthesised antigen-active peptides, imitating them. Some regularities of structural and functional organization of adenovirus hexon have been determined on the basis of comparative analysis of antigenic specificity and primary structure of proteins being apart in taxonomic respect. The conception of immunoactivation (infectivity neutralization) of adenoviruses has been developed, and a mathematical model of this process has been first proposed, which determines the impact of antibodies to several antigenic determinants of hexon and fiber as well as interferon and complement. The unknown peculiarities of the adenovirus genome expression were studied in the dynamics of productive infection or under the effect of modified nucleosides, proteolysis inhibitors and those of different nature promising for chemotherapy of adenovirus infection. Lymphotropicity of adenoviruses was established and a model of the mixed infection of lymphocytes with adenoviruses, HIV, and Epstein-Barr virus of the herpes virus family was proposed for the first time. It was determined that the mutual interference of viruses was developed at the process of a single or successive infection and this was important to understand AIDS immunopathogenesis. Data presented substantiate the ways of creation of modern efficient preparations for diagnosis, prophylaxis and chemotherapy of adenovirus infection.

[The viral genome status studied under the conditions of a mixed infection in lymphoblastoid cells by adenovirus and the Epstein-Barr virus]. / Izuchenie sostoianiia virusnykh genomov v usloviiakh smeshannoi infektsii limfoblastoidnykh kletok adenovirusom i virusom Epshteina-Barr.

Some indices have been studied which characterized the state of Epstein-Barr virus genome and adenovirus in the implanted lines of lymphoblastoid cells of B and T phenotype under the mixed or monoinfection. It has been shown that super infection by type 2 adenovirus rather sharply affects the state of Epstein-Barr virus genome in the Raji cells containing integrated Epstein-Barr virus genome. The state of adenovirus genome in the studied cells is less subject to changes. Its early area is revealed by hybridization using DNA-DNA method in a form of two fragments of different intensity which is maximum in the Raji and Jurkat cells, which evidences for the more expressivity of adenovirus genome in these cells.