Leptolysin, a Leptospira secreted metalloprotease of the pappalysin family with broad-spectrum activity

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Courrol, Daniella dos Santos ; Da Silva, Cristiane Castilho Fernandes ; Prado, Luan Gaviao ; Chura-Chambi, Rosa Maria ; Morganti, Ligia ; De Souza, Gisele Oliveira ; Heinemann, Marcos Bryan ; Isaac, Lourdes ; Conte, Fernando Paiva ; Vieira Portaro, Fernanda Calheta ; Rodrigues-da-Silva, Rodrigo Nunes ; Barbosa, Angela Silva

• Tipo de documento: Artigo Científico
• Fonte: FRONTIERS IN CELLULAR AND INFECTION MICROBIOLOGY; v. 12, p. 20-pg., 2022-08-23.

Resumo

Extracellular proteolytic enzymes are produced by a variety of pathogenic microorganisms, and contribute to host colonization by modulating virulence. Here, we present a first characterization of leptolysin, a Leptospira metalloprotease of the pappalysin family identified in a previous exoproteomic study. Comparative molecular analysis of leptolysin with two other pappalysins from prokaryotes, ulilysin and mirolysin, reveals similarities regarding calcium, zinc, and arginine -binding sites conservation within the catalytic domain, but also discloses peculiarities. Variations observed in the primary and tertiary structures may reflect differences in primary specificities. Purified recombinant leptolysin of L. interrogans was obtained as a similar to 50 kDa protein. The protease exhibited maximal activity at pH 8.0 and 37 degrees C, and hydrolytic activity was observed in the presence of different salts with maximum efficiency in NaCl. Substrate specificity was assessed using a small number of FRET peptides, and showed a marked preference for arginine residues at the P1 position. L. interrogans leptolysin proteolytic activity on proteinaceous substrates such as proteoglycans and plasma fibronectin was also evaluated. All proteins tested were efficiently degraded over time, confirming the protease' s broad-spectrum activity in vitro. In addition, leptolysin induced morphological alterations on HK-2 cells, which may be partially attributed to extracellular matrix (ECM) degradation. Hemorrhagic foci were observed in the dorsal skin of mice intradermally injected with leptolysin, as a plausible consequence of ECM disarray and vascular endothelium glycocalyx damage. Assuming that leptospiral proteases play an important role in all stages of the infectious process, characterizing their functional properties, substrates and mechanisms of action is of great importance for therapeutic purposes. (AU)

• Processo FAPESP: 18/12896-2 - Avaliação do papel de proteases secretadas por leptospira na degradação e inativação de moléculas do hospedeiro
• Beneficiário: Angela Silva Barbosa
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 19/22706-9 - Produção e caracterização da protease papalisina de Leptospira interrogans
• Beneficiário: Daniella dos Santos Courrol
• Modalidade de apoio: Bolsas no Brasil - Doutorado Direto
• Processo FAPESP: 17/12924-3 - Etiopatogênese da Leptospirose: contribuição do sistema complemento in vivo e in vitro para o controle da infecção e desencadeamento da resposta inflamatória tecidual: estudo de polimorfismos de genes do sistema complemento em pacientes com Leptospirose
• Beneficiário: Lourdes Isaac
• Modalidade de apoio: Auxílio à Pesquisa - Temático