RESUMO
Herpesviruses are large, ubiquitous DNA viruses with complex host interactions, yet many of the proteins encoded by these viruses have not been functionally characterized. As a first step in functional characterization, we determined the subcellular localization of 234 epitope-tagged proteins from herpes simplex virus, cytomegalovirus, and Epstein-Barr virus. Twenty-four of the 93 proteins with nuclear localization formed subnuclear structures. Twelve of these localized to the nucleolus, and five at least partially localized with promyelocytic leukemia (PML) bodies, which are known to suppress viral lytic infection. In addition, two proteins disrupted Cajal bodies, and 19 of the nuclear proteins significantly decreased the number of PML bodies per cell, including six that were shown to be SUMO-modified. These results have provided the first functional insights into over 120 previously unstudied proteins and suggest that herpesviruses employ multiple strategies for manipulating nuclear bodies that control key cellular processes.
Assuntos
Citomegalovirus/metabolismo , Herpesvirus Humano 4/metabolismo , Proteínas Nucleares/metabolismo , Simplexvirus/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Proteínas Virais/metabolismo , Estruturas do Núcleo Celular/genética , Estruturas do Núcleo Celular/metabolismo , Citomegalovirus/genética , Testes Genéticos , Genoma , Células Precursoras de Granulócitos/metabolismo , Herpesvirus Humano 4/genética , Humanos , Proteínas Nucleares/genética , Proteína da Leucemia Promielocítica , Simplexvirus/genética , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/genética , Replicação Viral , Dedos de ZincoRESUMO
Spiropiperidine naphthyridinone inhibitors of Staphylococcus aureus and Escherichia coli FabI have been prepared. Compounds 14a and 14c were identified as having sub-nanomolar E. coli FabI activity and are among the most potent FabI inhibitors yet described. The structural model of 14a bound to E. coli FabI is shown.
Assuntos
Antibacterianos/farmacologia , Enoil-(Proteína de Transporte de Acila) Redutase (NADH)/antagonistas & inibidores , Enoil-(Proteína de Transporte de Acila) Redutase (NADH)/metabolismo , Escherichia coli/enzimologia , Naftiridinas/farmacologia , Piperidinas/farmacologia , Compostos de Espiro/farmacologia , Staphylococcus aureus/enzimologia , Antibacterianos/química , Domínio Catalítico , Enoil-(Proteína de Transporte de Acila) Redutase (NADH)/química , Modelos Moleculares , Naftiridinas/química , Piperidinas/química , Ligação Proteica , Compostos de Espiro/química , Relação Estrutura-AtividadeRESUMO
In the search for new antibacterial agents, the enzyme FabI has been identified as an attractive target. Employing a structure guided approach, the previously reported ene-amide series of FabI inhibitors were expanded to include 2,3,4,5-tetrahydro-1H-pyrido[2,3-b and e][1,4]diazepines. These novel series incorporate additional H-bonding functions and can be more water soluble than their naphthyridinone progenitors; diazepine 16c is shown to be efficacious in a mouse infection model.
Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Azepinas/química , Azepinas/farmacologia , Enoil-(Proteína de Transporte de Acila) Redutase (NADH)/antagonistas & inibidores , Enoil-(Proteína de Transporte de Acila) Redutase (NADH)/metabolismo , Animais , Antibacterianos/uso terapêutico , Azepinas/uso terapêutico , Escherichia coli/efeitos dos fármacos , Escherichia coli/enzimologia , Infecções por Escherichia coli/tratamento farmacológico , Camundongos , Modelos Moleculares , Ligação Proteica , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/enzimologiaRESUMO
We have developed a high-throughput system for generating baculoviruses and testing the expression, solubility, and affinity column purification of encoded proteins. We have used this system to generate baculoviruses for and analyze the expression of 337 proteins from three different herpesviruses (HSV-1, EBV, and CMV) and vaccinia virus. Subsets of these proteins were also tested for expression and solubility in E. coli. Comparisons of the results in the two systems are presented for each virus.