Batroxase, a new metalloproteinase from B. atrox snake venom with strong fibrinolytic activity

• DOI: 10.1016/j.toxicon.2012.03.018
• Publication Date: 2012-03-30T08:43:50Z
• Authors: A.C.O. Cintra; L.G.B. De Toni; M.A. Sartim; J.J. Franco; R.C. Caetano; M.T. Murakami; S.V. Sampaio
• Keywords: Models, Molecular; 0303 health sciences; Sequence Homology, Amino Acid; Fibrinolysis; Molecular Sequence Data; Toxicology; Mass Spectrometry; 03 medical and health sciences; Crotalid Venoms; Metalloproteases; Animals; Bothrops; Amino Acid Sequence; Isoelectric Point

The structures and functional activities of metalloproteinases from snake venoms have been widely studied because of the importance of these molecules in envenomation. Batroxase, which is a metalloproteinase isolated from Bothrops atrox (Pará) snake venom, was obtained by gel filtration and anion exchange chromatography. The enzyme is a single protein chain composed of 202 amino acid residues with a molecular mass of 22.9 kDa, as determined by mass spectrometry analysis, showing an isoelectric point of 7.5. The primary sequence analysis indicates that the proteinase contains a zinc ligand motif (HELGHNLGISH) and a sequence C₁₆₄ I₁₆₅M₁₆₆ motif that is associated with a "Met-turn" structure. The protein lacks N-glycosylation sites and contains seven half cystine residues, six of which are conserved as pairs to form disulfide bridges. The three-dimensional structure of Batroxase was modeled based on the crystal structure of BmooMPα-I from Bothrops moojeni. The model revealed that the zinc binding site has a high structural similarity to the binding site of other metalloproteinases. Batroxase presented weak hemorrhagic activity, with a MHD of 10 μg, and was able to hydrolyze extracellular matrix components, such as type IV collagen and fibronectin. The toxin cleaves both α and β-chains of the fibrinogen molecule, and it can be inhibited by EDTA, EGTA and β-mercaptoethanol. Batroxase was able to dissolve fibrin clots independently of plasminogen activation. These results demonstrate that Batroxase is a zinc-dependent hemorrhagic metalloproteinase with fibrin(ogen)olytic and thrombolytic activity.