RESUMEN
The antigenic structure of the comovirus bean pod mottle virus (BPMV) was studied using synthetic peptides selected on the basis of the exposed location of certain regions of the viral protein. Three regions of domain A, four regions of domain B and two regions of domain C of BPMV coat protein were studied. Each of four regions were synthesized in the form of linear and cyclized peptides while the others were synthesized as linear peptides only. The peptides were tested for their ability to be recognized by antibodies directed against BPMV. The peptides were also used for producing rabbit antisera, which were tested for their ability to react with various BPMV antigens as well as with the linear and cyclized peptides. All the peptides were found to correspond to epitopes of BPMV coat protein. Several of the antigenic sites of BPMV located on exposed loops of the coat protein occupy positions which correspond to known epitopes in the structurally related picornaviruses. Only in some cases did cyclization sufficiently improve the level of conformational mimicry between peptides and the viral protein to allow cross-reactions between them to be observed.
Asunto(s)
Antígenos Virales/análisis , Cápside/inmunología , Virus de Plantas/inmunología , Secuencia de Aminoácidos , Animales , Antígenos Virales/química , Cápside/química , Pollos , Reacciones Cruzadas , Ensayo de Inmunoadsorción Enzimática , Epítopos/análisis , Epítopos/química , Sueros Inmunes/inmunología , Datos de Secuencia Molecular , Fragmentos de Péptidos/síntesis química , Fragmentos de Péptidos/química , Fragmentos de Péptidos/inmunología , Estructura Terciaria de Proteína , ConejosAsunto(s)
Inmunoensayo/métodos , Proteínas/análisis , Animales , Anticuerpos Monoclonales , Antígenos/análisis , Reacciones Cruzadas , Ensayo de Inmunoadsorción Enzimática/instrumentación , Ensayo de Inmunoadsorción Enzimática/métodos , Immunoblotting/métodos , Inmunodifusión/métodos , Indicadores y Reactivos , Pruebas de Precipitina , Proteínas/genética , Proteínas/inmunología , Proteínas Virales/análisisRESUMEN
Monoclonal antibodies (mAb) capable of reacting with different potyviruses were obtained by immunizing mice with proteolysed tobacco etch virus. The mAb were not equally effective in all ELISA formats and some were specific for different conformational states of the viral coat protein. The mAb also detected antigenic differences between purified virus particles and viral antigen in infected plant sap. In an ELISA format using antigen-coated plates, 5 different potyviruses (out of 7 viruses tested) could be detected in plant sap by one mAb. Different combinations of mAb and polyclonal antiserum could also be used for detecting several potyviruses by ELISA.
Asunto(s)
Anticuerpos Monoclonales/química , Reacciones Cruzadas , Nicotiana/microbiología , Virus de Plantas/inmunología , Plantas Tóxicas , Secuencia de Aminoácidos , Anticuerpos Monoclonales/biosíntesis , Cápside/síntesis química , Cápside/inmunología , Ensayo de Inmunoadsorción Enzimática , Hibridomas/metabolismo , Datos de Secuencia Molecular , Fragmentos de Péptidos/inmunología , Enfermedades de las Plantas/microbiología , Extractos Vegetales/inmunología , Virus de Plantas/químicaRESUMEN
Eight peptides corresponding to conserved regions of the coat protein of potyviruses were synthesized. All the peptides were recognized by anti-virus or anti-core-virus. Antisera raised to the synthetic peptides were tested with purified viruses and viral antigens present in plant sap. In many cases, the extent of cross-reactivity between different potyviruses was not correlated with the degree of sequence homology between the peptide used for immunization and the corresponding region in the coat protein of the potyvirus tested. An antiserum raised to a peptide of 18 residues containing a highly conserved region was found to react with all seven potyviruses tested.
Asunto(s)
Sueros Inmunes/química , Fragmentos de Péptidos/inmunología , Virus de Plantas/aislamiento & purificación , Secuencia de Aminoácidos , Animales , Reacciones Antígeno-Anticuerpo , Pollos , Ensayo de Inmunoadsorción Enzimática , Epítopos/química , Epítopos/inmunología , Datos de Secuencia Molecular , Fragmentos de Péptidos/síntesis química , Enfermedades de las Plantas/microbiología , Extractos Vegetales/inmunología , Virus de Plantas/inmunología , Conejos , Proteínas del Núcleo Viral/inmunologíaRESUMEN
Middle component particles of bean pod mottle virus (BPMV) containing small protein subunits with a cleaved C terminus were used to produce monoclonal antibodies (MAbs). All MAbs were specific for cryptotopes, i.e. epitopes present only on dissociated BPMV protein. The MAbs reacted more strongly with virus protein preparations containing the cleaved form of the small subunit than with preparations containing only the uncleaved form. It seems that the presence of additional residues at the C terminus of the intact small subunit interferes with antibody binding. Antibodies raised against synthetic peptides corresponding to the C terminus of the uncleaved small subunit reacted with both intact virions and dissociated subunits. This C-terminal region seems to play a dominant role in the antigenicity of the virus.