Purification and functional characterization of two fibrinogenolytic enzymes from Bothrops alternatus venom

Two fibrinogenolytic enzymes, Bothrops alternatus metalloprotease isoform (BaltMP)-I and II, were purified from Bothrops alternatus venom using Diethylaminoethyl (DEAE) Sephacel, Sephadex G-75 and Heparin-Agarose column chromatography. Purified BaltMP-I and II ran as single protein bands on analytic...

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Bibliographic Details
Published in:Journal of Venomous Animals and Toxins including Tropical Diseases
Main Authors: J. O. Costa, C. B. Petric, A. Hamaguchi, M. I. Homsi-Brandeburgo, C. Z. Oliveira, A. M. Soares, F. Oliveira
Format: Article in Journal/Newspaper
Language:English
Published: SciELO 2007
Subjects:
snake venom
Bothrops alternatus
metalloproteases
functional characterization
fibrinogenolytic activity
defibrinogenation in vivo
Arctic medicine. Tropical medicine
RC955-962
Toxicology. Poisons
RA1190-1270
Zoology
QL1-991
Arctic
Online Access:https://doi.org/10.1590/S1678-91992007000300007
https://doaj.org/article/6fb72fe992b64298915b9eb097080460
Description
Summary:Two fibrinogenolytic enzymes, Bothrops alternatus metalloprotease isoform (BaltMP)-I and II, were purified from Bothrops alternatus venom using Diethylaminoethyl (DEAE) Sephacel, Sephadex G-75 and Heparin-Agarose column chromatography. Purified BaltMP-I and II ran as single protein bands on analytical polyacrylamide gel electrophoresis and showed molecular weights of 29000 and 36000, respectively, under reducing conditions in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). BaltMP-II, but not BaltMP-I, displayed blood-clotting activity in bovine plasma, which was about 10-fold higher than that of the crude venom. Both enzymes were proteolytically active against bovine fibrinogen as substrate. When fibrinogen and each enzyme were incubated at 37°C, at a ratio of 1:100 (w/w), BaltMP-II cleaved preferentially the Aalpha -chain and more slowly the Bbeta -chain. The action of BaltMP-I was similar, but lower. None of the proteases degraded the gamma-chain of fibrinogen. The fibrinogenolytic activity of the enzymes was inhibited by 1,10-phenanthroline, suggesting they are metalloproteases. Since both enzymes were found to cause defibrinogenation when intraperitoneally (i.p.) administered to mice, they can be of medical interest as a therapeutic agent in the treatment and prevention of arterial thrombosis.

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