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Adhesins of Leptospira interrogans mediate the interaction to fibrinogen and inhibit fibrin clot formation in vitro.

Oliveira R, Domingos RF, Siqueira GH, Fernandes LG, Souza NM, Vieira ML, de Morais ZM, Vasconcellos SA, Nascimento AL - PLoS Negl Trop Dis (2013)

Bottom Line: The data suggest that PLA-coated leptospires were capable to employ their proteolytic activity to decrease one substrate of the coagulation cascade.We also present six leptospiral adhesins and PLG- interacting proteins, rLIC12238, Lsa33, Lsa30, OmpL1, rLIC11360 and rLIC11975, as novel Fg-binding proteins.The recombinant proteins interact with Fg in a dose-dependent and saturable fashion when increasing protein concentration was set to react to a fix human Fg concentration.

View Article: PubMed Central - PubMed

Affiliation: Centro de Biotecnologia, Instituto Butantan, São Paulo, São Paulo, Brazil.

ABSTRACT
We report in this work that Leptospira strains, virulent L. interrogans serovar Copenhageni, attenuated L. interrogans serovar Copenhageni and saprophytic L. biflexa serovar Patoc are capable of binding fibrinogen (Fg). The interaction of leptospires with Fg inhibits thrombin- induced fibrin clot formation that may affect the haemostatic equilibrium. Additionally, we show that plasminogen (PLG)/plasmin (PLA) generation on the surface of Leptospira causes degradation of human Fg. The data suggest that PLA-coated leptospires were capable to employ their proteolytic activity to decrease one substrate of the coagulation cascade. We also present six leptospiral adhesins and PLG- interacting proteins, rLIC12238, Lsa33, Lsa30, OmpL1, rLIC11360 and rLIC11975, as novel Fg-binding proteins. The recombinant proteins interact with Fg in a dose-dependent and saturable fashion when increasing protein concentration was set to react to a fix human Fg concentration. The calculated dissociation equilibrium constants (K D ) of these reactions ranged from 733.3 ± 276.8 to 128 ± 89.9 nM for rLIC12238 and Lsa33, respectively. The interaction of recombinant proteins with human Fg resulted in inhibition of fibrin clot by thrombin-catalyzed reaction, suggesting that these versatile proteins could mediate Fg interaction in Leptospira. Our data reveal for the first time the inhibition of fibrin clot by Leptospira spp. and presents adhesins that could mediate these interactions. Decreasing fibrin clot would cause an imbalance of the coagulation cascade that may facilitate bleeding and help bacteria dissemination.

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Related in: MedlinePlus

Recognition of Fg binding to Leptopsira by IFA.Live L. interrogans sorovar Copenhageni strain M-20 were fixed with 2% paraformaldehyde and treated wit 16 µg of Fg. The recognition was assessed through polyclonal anti-Fg antibodies under confocal immunofluorescence microscope (Fg1 and Fg2). Control in which Fg was absent from the incubation mixture is shown (Fg Ø). Panels A: DNA propidium iodide-stained, B: FITC-stained and C: A+B composite images.
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pntd-0002396-g001: Recognition of Fg binding to Leptopsira by IFA.Live L. interrogans sorovar Copenhageni strain M-20 were fixed with 2% paraformaldehyde and treated wit 16 µg of Fg. The recognition was assessed through polyclonal anti-Fg antibodies under confocal immunofluorescence microscope (Fg1 and Fg2). Control in which Fg was absent from the incubation mixture is shown (Fg Ø). Panels A: DNA propidium iodide-stained, B: FITC-stained and C: A+B composite images.

Mentions: The ability of L. interrogans sorovar Copenhageni strain M-20 cells to bind human Fg was performed by immunofluorescence assay (IFA). Leptospires were visualized by propidium iodide staining (Fig. 1, panel A) followed by protein detection with goat anti- human Fg, in the presence of anti-goat IgG antibodies conjugated to FITC. Green fluorescence could be observed for Fg (Fig. 1 -Fg1B, Fg2B). The localization of the protein-green light within the leptospires was achieved by superimposing both fields and the results obtained are shown in Fig. 1 - Fg1C and Fg2C. The FgØ shows the control of the reaction in which Fg was absent.


Adhesins of Leptospira interrogans mediate the interaction to fibrinogen and inhibit fibrin clot formation in vitro.

Oliveira R, Domingos RF, Siqueira GH, Fernandes LG, Souza NM, Vieira ML, de Morais ZM, Vasconcellos SA, Nascimento AL - PLoS Negl Trop Dis (2013)

Recognition of Fg binding to Leptopsira by IFA.Live L. interrogans sorovar Copenhageni strain M-20 were fixed with 2% paraformaldehyde and treated wit 16 µg of Fg. The recognition was assessed through polyclonal anti-Fg antibodies under confocal immunofluorescence microscope (Fg1 and Fg2). Control in which Fg was absent from the incubation mixture is shown (Fg Ø). Panels A: DNA propidium iodide-stained, B: FITC-stained and C: A+B composite images.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3757074&req=5

pntd-0002396-g001: Recognition of Fg binding to Leptopsira by IFA.Live L. interrogans sorovar Copenhageni strain M-20 were fixed with 2% paraformaldehyde and treated wit 16 µg of Fg. The recognition was assessed through polyclonal anti-Fg antibodies under confocal immunofluorescence microscope (Fg1 and Fg2). Control in which Fg was absent from the incubation mixture is shown (Fg Ø). Panels A: DNA propidium iodide-stained, B: FITC-stained and C: A+B composite images.
Mentions: The ability of L. interrogans sorovar Copenhageni strain M-20 cells to bind human Fg was performed by immunofluorescence assay (IFA). Leptospires were visualized by propidium iodide staining (Fig. 1, panel A) followed by protein detection with goat anti- human Fg, in the presence of anti-goat IgG antibodies conjugated to FITC. Green fluorescence could be observed for Fg (Fig. 1 -Fg1B, Fg2B). The localization of the protein-green light within the leptospires was achieved by superimposing both fields and the results obtained are shown in Fig. 1 - Fg1C and Fg2C. The FgØ shows the control of the reaction in which Fg was absent.

Bottom Line: The data suggest that PLA-coated leptospires were capable to employ their proteolytic activity to decrease one substrate of the coagulation cascade.We also present six leptospiral adhesins and PLG- interacting proteins, rLIC12238, Lsa33, Lsa30, OmpL1, rLIC11360 and rLIC11975, as novel Fg-binding proteins.The recombinant proteins interact with Fg in a dose-dependent and saturable fashion when increasing protein concentration was set to react to a fix human Fg concentration.

View Article: PubMed Central - PubMed

Affiliation: Centro de Biotecnologia, Instituto Butantan, São Paulo, São Paulo, Brazil.

ABSTRACT
We report in this work that Leptospira strains, virulent L. interrogans serovar Copenhageni, attenuated L. interrogans serovar Copenhageni and saprophytic L. biflexa serovar Patoc are capable of binding fibrinogen (Fg). The interaction of leptospires with Fg inhibits thrombin- induced fibrin clot formation that may affect the haemostatic equilibrium. Additionally, we show that plasminogen (PLG)/plasmin (PLA) generation on the surface of Leptospira causes degradation of human Fg. The data suggest that PLA-coated leptospires were capable to employ their proteolytic activity to decrease one substrate of the coagulation cascade. We also present six leptospiral adhesins and PLG- interacting proteins, rLIC12238, Lsa33, Lsa30, OmpL1, rLIC11360 and rLIC11975, as novel Fg-binding proteins. The recombinant proteins interact with Fg in a dose-dependent and saturable fashion when increasing protein concentration was set to react to a fix human Fg concentration. The calculated dissociation equilibrium constants (K D ) of these reactions ranged from 733.3 ± 276.8 to 128 ± 89.9 nM for rLIC12238 and Lsa33, respectively. The interaction of recombinant proteins with human Fg resulted in inhibition of fibrin clot by thrombin-catalyzed reaction, suggesting that these versatile proteins could mediate Fg interaction in Leptospira. Our data reveal for the first time the inhibition of fibrin clot by Leptospira spp. and presents adhesins that could mediate these interactions. Decreasing fibrin clot would cause an imbalance of the coagulation cascade that may facilitate bleeding and help bacteria dissemination.

Show MeSH
Related in: MedlinePlus