In vitro effects of Echis carinatus venom on the human plasma proteome

Echis carinatus venom (EV) is a complex mixture of toxins that contribute to its lethality. EV proteolytic activity was analyzed by zymography, chromogenic assays, and SDS-PAGE. To understand the molecular mechanism of the envenomation, we investigated the in vitro effect of EV on human plasma prote...

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Bibliographic Details
Published in:Proteomics (Weinheim) 2010-10, Vol.10 (20), p.3712-3722
Main Authors: Guerranti, Roberto, Cortelazzo, Alessio, Hope-Onyekwere, Nnadozie S, Furlani, Emiliano, Cerutti, Helena, Puglia, Michele, Bini, Luca, Leoncini, Roberto
Format: Article
Language:English
Subjects:
2-D PAGE
Analytical, structural and metabolic biochemistry
Animal proteomics
Animals
Biological and medical sciences
Blood Proteins - chemistry
Blood Proteins - metabolism
Echis carinatus
Electrophoresis, Polyacrylamide Gel - methods
Fundamental and applied biological sciences. Psychology
Humans
Miscellaneous
Molecular Sequence Data
Peptides - analysis
Plasma proteome
Proteases
Protein fragmentation
Proteins
Proteome - analysis
Proteome - drug effects
Snake venom
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - methods
Tandem Mass Spectrometry - methods
Viper Venoms - chemistry
Viper Venoms - pharmacology
Viperidae
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Summary:Echis carinatus venom (EV) is a complex mixture of toxins that contribute to its lethality. EV proteolytic activity was analyzed by zymography, chromogenic assays, and SDS-PAGE. To understand the molecular mechanism of the envenomation, we investigated the in vitro effect of EV on human plasma proteins. We looked for EV protein substrates and their proteolytic fragments. We analyzed EV proteolytic activity on standard proteins such as prothrombin or fibrinogen. To set up the optimal EV:plasma protein ratio conditions, plasma was incubated with EV (treated plasma), depleted of abundant proteins, and subjected to SDS-PAGE. Samples from control and treated plasma were also analyzed by 2-DE/MALDI-TOF MS, leading to the identification of four classes of plasma proteins cleaved by EV: proteases, protease inhibitors, binding proteins, and transporters. EV mainly proteolyzes entire proteins but can also act on physiological fragments. In summary, the physiological effects of EV proteases involve other important processes in addition to blood coagulation; complement activation and hemoglobin metabolism are also affected. In particular, the cleavage of protease inhibitors appears to be the mechanism through which the venom neutralizes the body's defenses.
ISSN:1615-9853
1615-9861
1615-9861
DOI:10.1002/pmic.201000055