Identificação de inibidores específicos para a protease NS2B-NS3 do flavivírus Zika

• Ano de defesa: 2020
• Autor(a) principal: Santo, Camila Cardoso Di [UNIFESP]
• Tipo de documento: Dissertação
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de São Paulo (UNIFESP)
• Palavras-chave em Português: Zika Vírus; Inibidores De Serinoproteases; Phage Display; Docking
• Link de acesso: https://sucupira.capes.gov.br/sucupira/public/consultas/coleta/trabalhoConclusao/viewTrabalhoConclusao.jsf?popup=true&id_trabalho=8473363; https://hdl.handle.net/11600/64284

Besides Dengue, Chikungunya and Yellow Fever Aedes aegypti mosquitoes also transmits Zika virus, and despite vigorous reasearch no vaccine or antiviral is available, and considering no therapy is available a specific Zika virus inhhibitor would allow a faster drug treatment. Zika does not present as severe symptoms as Dengue or Chikungunya, but cases of microcephaly and Guillan-Barré syndrome have been reported, specially microcephaly in fetuses and newborn child, and both consequences of utmost importance. Among viral proteases, NS2B-NS3 represents one of the most studied drug targets due to its role in viral replication, cleaving flaviviral poliproteins into structural and non-structural proteins. In order to find such a specific inhibitor the present work focused on two approaches: 1) identify an inhibitor through kinetic assays of previously characterized molecules from Laboratório de Bioquímica e Biologia Molecular de Artrópodes e Hematófagos (UNIFESP-EPM); and 2) through phage display technique expose the purified recombinant NS2B-NS3 protein to a mutant library previously elaborated. This library, TiPI 1, carry mutations from P2 to P2’, including its reactive site. Regarding the screening, two inhibitors were found: 1) BmKK with Ki of 36.01 nM and 2) Boophilin D1 with Ki of 15.53 nM, in which docking studies suggests interaction between residues D83 from NS2B and reactive site residue K31 from Boophilin D1, the D83 residue was previously described as important for correct folding and therefore activity of NS3 protease. Regarding phage display technique, no unique and redundant mutant sequence was obtained and therefore explored as possible inhibitor, glutamine residues were found throughout the sequences, in P2, P1, P1’ and P2’ positions. Besides no positive results regarding TiPI 1 a new mutant library with the best inhibitor founded, Boophilin D1, should be studied in order to enhance the pursuit of a specific Zika virus inhibitor, enabling even the applications of this method to other flaviviral proteases.