ABSTRACT
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.
Subject(s)
Plant Diseases/virology , Rhabdoviridae/metabolism , Ribonucleoproteins/metabolism , Solanum tuberosum , Viral Proteins/biosynthesis , Cell Membrane/metabolism , Protein BiosynthesisABSTRACT
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.
Subject(s)
Plant Diseases/virology , RNA-Dependent RNA Polymerase/metabolism , Rhabdoviridae/genetics , Ribonucleoproteins/metabolism , Solanum tuberosum/virology , Viral Proteins/metabolism , Electrophoresis, Polyacrylamide Gel , Polyribosomes/genetics , Polyribosomes/metabolism , RNA, Viral/analysis , RNA, Viral/biosynthesis , RNA-Dependent RNA Polymerase/genetics , Rhabdoviridae/metabolism , Ribonucleoproteins/genetics , Viral Proteins/genetics , Viral Proteins/pharmacology , Virus Replication/drug effects , Virus Replication/geneticsABSTRACT
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.
Subject(s)
Antigens/immunology , Nucleocapsid Proteins/immunology , Plant Viruses/immunology , Rabies virus/immunology , Vesicular stomatitis Indiana virus/immunology , Electrophoresis, Polyacrylamide Gel , Immunoblotting , Immunodiffusion , Molecular Weight , Nucleocapsid Proteins/chemistry , Nucleocapsid Proteins/genetics , Nucleocapsid Proteins/isolation & purification , Plant Viruses/genetics , Rabies virus/genetics , Vesicular stomatitis Indiana virus/geneticsABSTRACT
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.
