RESUMO
Mab7.3 to Yersinia pestis LcrV antigen (LcrV(Ype)) protected J774A.1 macrophages in vitro from killing by a Yersinia pseudotuberculosis strain expressing LcrV(Ype). Of 4 site-directed mutations in the coiled-coil region (148-169) and 7 mutations in the 225-255 sequence of LcrV(Ype), only the mutation of N255 to D255, abrogated the binding of Mab7.3 and reduced its protective capacity against plague. Since the Mab7.3 epitope in LcrV(Ype) (135-275) encompasses a region (136-180) thought to be exposed on the injectisome, we suggest that Mab7.3 protects by binding to LcrV(Ype) and interfering with protein-protein interactions necessary for type three secretion.
Assuntos
Anticorpos Antibacterianos/imunologia , Anticorpos Monoclonais/imunologia , Antígenos de Bactérias/imunologia , Peste/prevenção & controle , Proteínas Citotóxicas Formadoras de Poros/imunologia , Sequência de Aminoácidos , Animais , Afinidade de Anticorpos , Linhagem Celular , Epitopos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Peste/imunologia , Vacina contra a Peste/imunologia , Estrutura Terciária de Proteína , Proteínas Recombinantes/imunologia , Yersinia pestis/imunologiaAssuntos
Proteínas de Bactérias/imunologia , Vacina contra a Peste , Peste/imunologia , Yersinia pestis/imunologia , Animais , Proteínas de Bactérias/genética , Humanos , Camundongos , Peste/prevenção & controle , Plasmídeos , Subunidades Proteicas/imunologia , Virulência , Yersinia pestis/genética , Yersinia pestis/patogenicidadeRESUMO
Monoclonal antibodies specific for Yersinia pestis V antigen and F1 antigen, administered singly or in combination, protected mice in models of bubonic and pneumonic plague. Antibodies showed synergy when administered prophylactically and as a therapy 48 h postinfection. Monoclonal antibodies therefore have potential as a treatment for plague.
Assuntos
Anticorpos Antibacterianos/administração & dosagem , Anticorpos Monoclonais/administração & dosagem , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Peste/prevenção & controle , Peste/terapia , Yersinia pestis/imunologia , Animais , Anticorpos Antibacterianos/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Modelos Animais de Doenças , Sinergismo Farmacológico , Quimioterapia Combinada , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Citotóxicas Formadoras de Poros , Yersinia pestis/patogenicidadeRESUMO
Live attenuated mutants of several pathogenic bacteria have been exploited as potential vaccine vectors for heterologous antigen delivery by the mucosal route. Such live vectors offer the advantage of potential delivery in a single oral, intranasal or inhalational dose, stimulating both systemic and mucosal immune responses. Over the years, a range of strategies have been developed to allow controlled and stable delivery of antigens and improved immunogenicity where required. Most of these approaches have been evaluated in Salmonella vaccine vectors and, as a result, several live attenuated recombinant Salmonella vaccines are now in human clinical trials. In this review, these strategies and their use in the development of a delivery system for the Yersinia pestis V antigen are described.
Assuntos
Antígenos de Bactérias/genética , Antígenos Heterófilos/genética , Vacinas Bacterianas/genética , Vacinas de DNA/administração & dosagem , Yersinia pestis/imunologia , Animais , Antígenos de Bactérias/imunologia , Antígenos Heterófilos/imunologia , Vacinas Bacterianas/administração & dosagem , Vacinas Bacterianas/imunologia , Humanos , Mucosa Bucal , Mucosa , Proteínas Citotóxicas Formadoras de Poros , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Vacinas de DNA/genética , Vacinas de DNA/imunologiaRESUMO
Attenuated strains of Salmonella enterica serovar Typhimurium are used as carriers of heterologous antigens as candidate oral vaccines and, more recently, as carriers of DNA vaccines. In this study, recombinant Salmonella strains that were altered in their ability to colonise murine tissues in vivo when compared to parent strains were not, however, equally altered in their ability to invade murine cells in vitro. These results suggest that in vitro invasion studies may not be a representative model for colonisation of tissues in vivo, and that in vitro studies should ideally be used in conjunction with in vivo studies for the assessment of potential Salmonella vaccines.
Assuntos
Vacinas Bacterianas/genética , Plasmídeos/genética , Salmonella typhimurium/genética , Salmonella typhimurium/imunologia , Animais , Antígenos/genética , Vacinas Bacterianas/imunologia , Linhagem Celular , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Recombinação Genética , Salmonella typhimurium/patogenicidade , Vacinas Tíficas-Paratíficas/genética , Vacinas Tíficas-Paratíficas/imunologia , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Vacinas de DNA/genética , Vacinas de DNA/imunologiaRESUMO
The type III secretion system is used by pathogenic Yersinia to translocate virulence factors into the host cell. A key component is the multifunctional LcrV protein, which is present on the bacterial surface prior to host cell contact and up-regulates translocation by blocking the repressive action of the LcrG protein on the cytosolic side of the secretion apparatus. The functions of LcrV are proposed to involve self-interactions (multimerization) and interactions with other proteins including LcrG. Coiled-coil motifs predicted to be present are thought to play a role in mediating these protein-protein interactions. We have purified recombinant LcrV, LcrG, and site-directed mutants of LcrV and demonstrated the structural integrity of these proteins using circular dichroism spectroscopy. We show that LcrV interacts both with itself and with LcrG and have obtained micromolar and nanomolar affinities for these interactions, respectively. The effects of LcrV mutations upon LcrG binding suggest that coiled-coil interactions indeed play a significant role in complex formation. In addition, comparisons of secretion patterns of effector proteins in Yersinia, arising from wild type and mutants of LcrV, support the proposed role of LcrG in titration of LcrV in vivo but also suggest that other factors may be involved.