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  • Enolases are among the most abundantly expressed


    Enolases are among the most abundantly expressed cytosolic proteins. They are metalloenzymes that catalyze the conversion of 2-phosphoglycerate to phosphoenolpyruvate [18]. Eubacteria and archaebacteria have a single enolase gene (reviewed in [19]). Despite the lack of signal peptides or cognate protein secretion systems, they have been found localized at the surface of a variety of eukaryotic and prokaryotic cells, where they display multiple moonlighting functions [20]. Enolase of both Gram-positive and Gram-negative bacteria has been shown to bind host plasminogen. Functional consequences of plasminogen binding by these microorganisms include dissemination and transmigration, evasion from host\'s innate immune responses and enhanced adhesion to host cells, as shown for Streptococcus pneumoniae, known to use surface-displayed enolase to interact with cell-surface-bound plasminogen [21]. Recent findings by Mori and colleagues have shown that S. pneumoniae α-enolase induces cell death of neutrophils by releasing neutrophil extracellular traps [22]. Moreover, pneumococcal enolase was shown to bind the negative complement regulator C4BP, thus contributing to complement evasion [23]. Several microorganisms also employ enolase as an adhesin. In Staphylococcus aureus α-enolase was identified as a laminin-binding protein [24], and surface-exposed α-enolases from Streptococcus suis, Lactobacillus plantarum and Paracoccidioides brasiliensis were characterized as fibronectin-binding proteins [25], [26], [27]. More recently, S. gallolyticus enolase has been shown to interact with a surface-exposed component of cytokeratin 8 (CK8) of human colonocytes, possibly contributing to bacterial adherence and infection [28]. Oral bacteria, including S. mutans and S. gordonii, also display α-enolase on the surface, and the salivary mucin MUC7 is one of the host\'s targets for this moonlighting protein. α-Enolase has also been shown to mediate adhesion of Mycoplasma suis to porcine erythrocytes, a crucial step in the life Cyt387 of this hemotrophic bacterium [29]. Recent published data have shown that enolase from Leptospira interrogans is secreted into the extracellular medium and then reassociates with the bacterial surface by interacting with outer membrane proteins [17]. Cyt387 Anti-enolase antibodies recognize intact, non-permeabilized leptospires, thus indicating that the protein is indeed surface-exposed. As shown for many other bacteria, membrane-anchored leptospiral enolase displays plasminogen binding activity [17]. Given the role of enolase in bacterial pathogenesis, we aimed to further characterize this multifunctional protein in Leptospira. In the present work, the functional consequences of plasminogen binding to leptospiral enolase were assessed, and the interaction of this particular protein with other host molecules was evaluated. Here we demonstrate that plasminogen bound to enolase is converted to plasmin, which in turn degrades natural substrates, including fibrinogen and vitronectin. Moreover, leptospiral enolase binds the complement regulators FH and C4BP. Once bound to enolase, both FH and C4BP remain functional, acting as cofactors for factor I (FI) in the cleavage of C3b and C4b, respectively. Taken together, our data suggest that enolase may contribute to leptospiral pathogenesis.
    Materials and methods
    Discussion Multiple requirements are necessary for successful infection by pathogens. In short, pathogenic microorganisms must be able to enter the host, remain in sites where the infection gets established and avoid or compromise host\'s immune defenses. To accomplish this program, pathogens have evolved mechanisms to overcome the natural host\'s barriers, among which are a range of virulence factors [39]. Many of them are surface-exposed molecules acting as adhesins, invasins and/or evasins. In addition to the panoply of outer membrane proteins having a role in microbial pathogenesis described to date, a growing number of moonlighting proteins has been reported over the last few years that are presumed to display diverse functions related to virulence and survival inside the host.