A Newly Identified Leptospiral Adhesin Mediates Attachment to Laminin

Abstract

Pathogenic leptospires have the ability to survive and disseminate to multiple organs after penetrating the host. Several pathogens, including spirochetes, have been shown to express surface proteins that interact with the extracellular matrix (ECM). This adhesin-mediated binding process seems to be a crucial step in the colonization of host tissues. This study examined the interaction of putative leptospiral outer membrane proteins with laminin, collagen type I, collagen type IV, cellular fibronectin, and plasma fibronectin. Six predicted coding sequences selected from the Leptospira interrogans serovar Copenhageni genome were cloned, and proteins were expressed, purified by metal affinity chromatography, and characterized by circular dichroism spectroscopy. Their capacity to mediate attachment to ECM components was evaluated by binding assays. We have identified a leptospiral protein encoded by LIC12906, named Lsa24 (leptospiral surface adhesin; 24 kDa) that binds strongly to laminin. Attachment of Lsa24 to laminin was specific, dose dependent, and saturable. Laminin oxidation by sodium metaperiodate reduced the protein-laminin interaction in a concentration-dependent manner, indicating that laminin sugar moieties are crucial for this interaction. Triton X-114-solubilized extract of L. interrogans and phase partitioning showed that Lsa24 was exclusively in the detergent phase, indicating that it is a component of the leptospiral membrane. Moreover, Lsa24 partially inhibited leptospiral adherence to immobilized laminin. This newly identified membrane protein may play a role in mediating adhesion of L. interrogans to the host. To our knowledge, this is the first leptospiral adhesin with laminin-binding properties reported to date.

Characterization of Three Novel Adhesins of Leptospira interrogans

“…[8][9][10][11][12][13][14][15][16][17][18] Several laminin-binding proteins were identified, which indicated that leptospires have a redundant repertoire of adhesion molecules that are probably part of their invasion strategies. Lsa23, Lsa26, and Lsa36 bind laminin in a dose-dependent, specific, and saturable manner, fulfilling the properties of receptor-ligand interaction.…”

“…Some of these proteins were identified as novel leptospiral adhesins that might be involved in the first steps of host-Leptospira interaction. [8][9][10][11][12][13][14][15][16][17][18] After attachment, leptospires have to overcome tissue barriers and reach target organs. Proteolytic activity by plasmin (PLA) generation at the surface of Leptospira spp.…”

Avaliação e caracterização de candidatos vacinais voltados para o controle da leptospirose.

“…Assim, a primeira proteína de leptospira com capacidade de ligação a laminina foi identificada e nomeada de Lsa24 (BARBOSA et al, 2006). A partir daí, diversas proteínas têm sido identificadas com capacidade de ligação a ECM (VIEIRA et al, 2014;.…”

“…Para isso, laminina, fibronectina celular, elastina, colágeno tipo I, colágeno tipo IV e as proteínas controles BSA e fetuína foram imobilizados em placas de microdiluição e a ligação das proteínas recombinantes foi avaliada por ELISA, após o uso dos anticorpos policlonais obtidos contra cada proteína recombinante. Várias proteínas de leptospiras com propriedades de ligação específica a laminina e a outros componentes da ECM foram identificadas pelo nosso grupo (ATZINGEN et al, 2008;ATZINGEN et al, 2009;BARBOSA et al, 2006;DOMINGOS et al, 2015;DOMINGOS et al, 2012;FERNANDES et al, 2014;FERNANDES et al, 2012;LONGHI et al, 2009;MENDES et al, 2011;OLIVEIRA et al, 2011;OLIVEIRA et al, 2010; __________________________________________________________Resultados e Discussão 113 2013; SOUZA et al, 2012;VIEIRA et al, 2010b). Outros grupos também identificaram proteínas com capacidade de adesão (CARVALHO et al, 2009;CHOY et al, 2007;HAUK et al, 2008;HOKE et al, 2008;LIN et al, 2009;STEVENSON et al, 2007).…”

“…(Figura 36). Nossos achados foram semelhantes aos resultados obtidos por Fernandes e colaboradores (2012) com a proteína OmpL1, mas, difere dos dados encontrados para a maioria dos outros ligantes de laminina (ATZINGEN et al, 2008;BARBOSA et al, 2006;DOMINGOS et al, 2012;OLIVEIRA et al, 2010). De qualquer forma, esses resultados indicam que nem todas as adesinas interagem com laminina na mesma região.…”