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Master's Dissertation
DOI
https://doi.org/10.11606/D.17.2021.tde-11062021-073458
Document
Author
Full name
Rafael de Souza Pontes
Institute/School/College
Knowledge Area
Date of Defense
Published
Ribeirão Preto, 2021
Supervisor
Committee
Stabeli, Rodrigo Guerino (President)
Beleboni, Renê de Oliveira
Oliveira, Arthur Henrique Cavalcante de
Title in Portuguese
Caracterização imunogênica do domínio B recombinante da proteína E2 de Chikungunya expressa em Escherichia coli
Keywords in Portuguese
Anticorpos
Arbovírus
CHIKV
Terapia de anticorpos
Abstract in Portuguese
O vírus Chikungunya (CHIKV) é o agente etiológico da febre Chikungunya (CHIKF), caracterizada por febre, erupção cutânea e artralgia. Parte dos indivíduos infectados progridem para artropatia crônica que pode persistir por anos. Nos últimos surtos foram reportadas complicações neurológicas, como encefalite, encefalopatia com alta mortalidade associada e síndrome de Guillain-Barré. No Brasil o vírus continua em circulação e em 2020 foram reportados mais de 60 mil casos e 14 óbitos. Apesar do interesse crescente nas pesquisas com CHIKV, não há nenhuma vacina ou tratamento específico disponível. O objetivo deste trabalho foi avaliar as propriedades imunogênicas do domínio B da proteína de envelope E2 de CHIKV expressa em sistema procarioto. Para isso, foram realizadas análises do grau de conservação das sequências de aminoácidos das proteínas de envelope do CHIKV e de epítopos de células B que são alvos de anticorpos neutralizantes expostos na superfície viral. As análises apontam para uma epítopo conservado localizado no domínio B da proteína E2 do CHIKV, que foi expressa utilizando a cepa Escherichia coli SHuflle. A estrutura secundária da proteína recombinante foi determinada por dicroísmo circular. Através de ensaio ELISA foi demonstrado a capacidade do soro de camundongos infectados com CHIKV de reconhecer o peptídeo recombinante. A imunização de camundongos Balb/c com o peptídeo recombinante elicitou altos títulos de anticorpos, que quando avaliados pelo teste de redução de plaque foram capazes de reduzir 50% das placas virais. Deste modo, o peptídeo recombinante expresso apresenta potencial para uso no desenvolvimento de vacinas, terapias baseadas em anticorpo e diagnóstico.
Title in English
Immunogenic characterization of the recombinant B domain of the Chikungunya E2 protein expressed in Escherichia coli
Keywords in English
Antibodies
Antibody therapy
Arbovirus
CHIKV
Abstract in English
Chikungunya virus (CHIKV) is the etiologic agent of Chikungunya fever (CHIKF), characterized by fever, rash, and arthralgia. Some infected individuals progress to chronic arthropathy, which can persist for years. In the last outbreaks, neurological complications have been reported, such as encephalitis, encephalopathy with high associated mortality, and Guillain-Barré syndrome. In Brazil, the virus remains in circulation, and in 2020 more than 60 thousand cases and 14 deaths were reported. Despite the growing interest in CHIKV research, there is no specific vaccine or treatment available. The aim of this work was to evaluate the immunogenic properties of domain B of the CHIKV E2 envelope protein expressed in a prokaryotic system. For that, analyzes of the degree of conservation of the amino acid sequences of the CHIKV envelope proteins and of B cell epitopes that are targets of neutralizing antibodies exposed on the viral surface were carried out. The analyzes point to a conserved epitope located in the B domain of the E2 protein of CHIKV, which was expressed using the strain Escherichia coli SHuflle. The secondary structure of the recombinant protein was determined by circular dichroism. The ELISA assay demonstrated the ability of serum from mice infected with CHIKV to recognize the recombinant peptide. The immunization of Balb / c mice with the recombinant peptide elicited high antibody titers, which when evaluated by the plaque reduction test were able to reduce 50% of the viral plaques. Thus, the expressed recombinant peptide has potential for use in the development of vaccines, antibody-based therapies, and diagnosis.
 
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Publishing Date
2021-06-18
 
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