Boophilin D1, a Kunitz type protease inhibitor, as a source of inhibitors for the ZIKA virus NS2B-NS3 protease

Autor(es)

Manzato, Veronica de Moraes ; Di Santo, Camila ; Torquato, Ricardo Jose Soares ; Coelho, Camila ; Gallo, Gloria ; Hardy, Leon ; Wurtele, Martin ; Tanaka, Aparecida Sadae

• Tipo de documento: Artigo Científico
• Fonte: Biochimie; v. 214, p. 6-pg., 2023-07-03.

Resumo

Arboviruses are a global concern for a multitude of reasons, including their increased incidence and human mortality. Vectors associated with arboviruses include the mosquito Aedes sp., which is responsible for transmitting the Zika virus. Flaviviruses, like the Zika virus, present only one chymotrypsin-like serine protease (NS3) in their genome. Together with host enzymes, the NS2B co-factor NS3 protease complex are essential for the viral replication cycle by virus polyprotein process-ing. To search for Zika virus NS2B-NS3 protease (ZIKVPro) inhibitors, a phage display library was con-structed using the Boophilin domain 1 (BoophD1), a thrombin inhibitor from the Kunitz family. A BoophilinD1 library mutated at positions P1-P40 was constructed, presenting a titer of 2.9x106 (cfu), and screened utilizing purified ZIKVPro. The results demonstrated at the P1-P4' positions the occurrence of 47% RALHA sequence (mut 12) and 11.8% RASWA sequence (mut14), SMRPT, or KALIP (wt) sequence. BoophD1-wt and mutants 12 and 14 were expressed and purified. The purified BoophD1 wt, mut 12 and 14, presented Ki values for ZIKVPro of 0.103, 0.116, and 0.101 mM, respectively. The BoophD1 mutant in-hibitors inhibit the Dengue virus 2 protease (DENV2) with Ki values of 0.298, 0.271, and 0.379 mM, respectively. In conclusion, BoophD1 mut 12 and 14 selected for ZIKVPro demonstrated inhibitory activity like BoophD1-wt, suggesting that these are the strongest Zika inhibitors present in the BoophD1 mutated phage display library. Furthermore, BoophD1 mutants selected for ZIKVPro inhibit both Zika and Dengue 2 proteases making them potential pan-flavivirus inhibitors.& COPY; 2023 Elsevier B.V. and Societe Francaise de Biochimie et Biologie Moleculaire (SFBBM). All rights reserved. (AU)

• Processo FAPESP: 12/03657-8 - Inibidores e proteases de ectoparasitas: relação de estrutura-função e identificação do papel dessas moléculas na interação de vetores de doenças e seus agentes etiológicos
• Beneficiário: Aparecida Sadae Tanaka
• Modalidade de apoio: Auxílio à Pesquisa - Temático
• Processo FAPESP: 19/03779-5 - Uso de phage display como ferramenta no diagnóstico e controle de doenças transmitidas por vetores hematófagos
• Beneficiário: Aparecida Sadae Tanaka
• Modalidade de apoio: Auxílio à Pesquisa - Temático