Autor(es)

De Carvalho, Daniella Romano ; Laurentino, Bruna Barbosa ; Rocha, Camila Loreta ; Kool, Jeroen ; Somsen, Govert ; van Hove, Erika Amstalden ; Cardoso, Carmen Lucia

• Tipo de documento: Artigo Científico
• Fonte: FRONTIERS IN ANALYTICAL SCIENCE; v. 2, p. 9-pg., 2022-10-19.

Resumo

Deregulated activity and expression of human kallikreins (KLKs) may be involved in various pathologies, so these enzymes are an attractive biological target for identifying molecules that can modulate KLK activity. This identification involves applying fast and efficient screening methods. This work describes an off-line assay with mass spectrometry (MS) detection that uses KLK immobilized on Sepharose-NHS as a micro-column configuration (IMER-KLK-Sepharose-NHS). The mass spectrometry used has an ion trap analyzer and electrospray ionization (EIS). The HPLC-MS method for quantifying KLK activity was developed. The enzymatic assay conditions were optimized, and the IMER-KLK-Sepharose-NHS kinetic parameter (KMapp = 15.48 +/- 3 mu mol L-1) was evaluated. Finally, the method was validated by using leupeptin as a reference inhibitor (IC50 = 0.85 +/- 0.10 mu mol L-1). The developed method was able to identify the reference inhibitor and can be an alternative for screening KLK inhibitors. (AU)

• Processo FAPESP: 14/50299-5 - EMU - espectrômetro de massas modelo Amazon speed com fonte ESI e duplo funil de íons, analisador tipo íon TRAP
• Beneficiário: Quezia Bezerra Cass
• Modalidade de apoio: Auxílio à Pesquisa - Programa Equipamentos Multiusuários
• Processo FAPESP: 14/50249-8 - Green chemistry: sustainable synthetic methods employing benign solvents, safer reagents, and bio-renewable feedstock
• Beneficiário: Arlene Gonçalves Corrêa
• Modalidade de apoio: Auxílio à Pesquisa - Programa Centros de Pesquisa Aplicada
• Processo FAPESP: 19/05363-0 - Triagem de ligantes enzimáticos: uma abordagem on-flow via LC-MS/MS
• Beneficiário: Carmen Lúcia Cardoso
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 16/14482-5 - Nanofracionamento por LC-MS/MS e ensaios at-line para a identificação de compostos biativos em extratos de produtos naturais.
• Beneficiário: Carmen Lúcia Cardoso
• Modalidade de apoio: Bolsas no Exterior - Pesquisa