Laboratório de Desenvolvimento de Vacinas, Instituto Butantan, 05503-900, São Paulo, Brazil; Programa de Pós-Graduação Interunidades em Biotecnologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, 05508-900, São Paulo, Brazil.
Laboratório de Bacteriologia, Instituto Butantan, 05503-900, São Paulo, Brazil.
Microbiol Res. 2020 May;235:126470. doi: 10.1016/j.micres.2020.126470. Epub 2020 Mar 19.
Leptospirosis is a global re-emerging zoonosis, caused by pathogenic bacteria of the genus Leptospira. Humans are infected mainly through contact with contaminated water or soil. The understanding of the molecular mechanisms of leptospirosis through the characterization of unknown outer membrane proteins may contribute to the development of new treatments, diagnostic methods and vaccines. We have identified using bioinformatics analysis a protein that is encoded by the gene LIC10774, predicted to be localized at the leptospiral outer membrane and exhibit beta-roll folding. Surface exposure was confirmed by flow cytometry, ELISA and immunofluorescence-based confocal microscopy. Through circular dichroism spectroscopy and hydrophobic dye binding we have shown that rLIC10774 binds calcium ions, which imposes changes to secondary and tertiary structures. The recombinant protein was capable of binding to several host extracellular matrix and serum components. Therefore, we describe LIC10774 as a calcium-binding protein exposed in the outer surface of pathogenic leptospires with possible multifunctional roles in adhesion to host tissues, evasion of the immune system and participation in dissemination processes during leptospirosis. In addition, we hypothesize that the calcium binding is important for temperature-dependent functional roles during leptospirosis.
钩端螺旋体病是一种全球性重新出现的动物传染病，由钩端螺旋体属的致病性细菌引起。人类主要通过接触受污染的水或土壤而感染。通过对未知外膜蛋白的特征描述来了解钩端螺旋体病的分子机制，可能有助于开发新的治疗方法、诊断方法和疫苗。我们通过生物信息学分析鉴定了一个由基因 LIC10774 编码的蛋白质，该蛋白预测定位于钩端螺旋体外膜，并呈现β-折叠结构。通过流式细胞术、ELISA 和基于免疫荧光的共聚焦显微镜证实了该蛋白的表面暴露。通过圆二色性光谱和疏水性染料结合实验，我们发现 rLIC10774 可以结合钙离子，从而导致二级和三级结构发生变化。重组蛋白能够与几种宿主细胞外基质和血清成分结合。因此，我们将 LIC10774 描述为一种暴露在致病性钩端螺旋体外表面的钙结合蛋白，它可能在与宿主组织的黏附、逃避免疫系统以及参与钩端螺旋体病的传播过程中具有多种功能。此外，我们假设钙结合对于钩端螺旋体病中温度依赖性的功能作用很重要。
