RESUMO
CMVpp65, a candidate component of human cytomegalovirus (CMV) vaccines, has phosphokinase (PK) activity that could affect vaccine safety. A mutated form of CMVpp65 substituting asparagine for lysine at the adenosine triphosphate (ATP)-binding site (CMVpp65mII) is kinase-deficient. Using DNA immunizations in a transgenic human leucocyte antigen (HLA)A*0201.Kb mouse model, the mutated CMVpp65 induced cytotoxic T lymphocytes (CTL) immunity similarly to native CMVpp65. Murine CTL lines generated from these immunizations killed human cells either after sensitization with CMVpp65-specific peptides or after infection with either CMV-Towne strain or rvac-pp65. It is proposed that CMVpp65mII be evaluated in candidate vaccines for CMV.
Assuntos
Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Antígenos HLA-A/imunologia , Fosfoproteínas/imunologia , Linfócitos T Citotóxicos/imunologia , Vacinas de DNA/imunologia , Proteínas da Matriz Viral/imunologia , Sequência de Aminoácidos , Animais , Células Cultivadas , Infecções por Citomegalovirus/prevenção & controle , Epitopos de Linfócito T/imunologia , Feminino , Humanos , Imunização , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Vacinas de DNA/genética , Vaccinia virus/genética , Proteínas da Matriz Viral/genética , Proteínas da Matriz Viral/metabolismoRESUMO
The major target of human cytomegalovirus (CMV)-specific cytotoxic T lymphocytes (CTL) is the tegument protein CMVpp65. However, this protein has protein kinase (PK) activity, and the unknown effects on cell replication of an exogenous PK in healthy cells could limit the use of CMVpp65 as a vaccine, especially in children. In this report we show that a point mutation converting lysine to asparagine at the invariant lysine (K436), an essential site for phosphotransfer, abolishes the threonine kinase activity. The mutant CMVpp65 maintains its immunologic target characteristics, including antibody and CTL reactivity. This kinase-deficient CMVpp65 is a candidate for evaluation in future CMV vaccine development.
