title: Structural and biochemical characterisation of VaF1, a P-IIIa fibrinogenolytic metalloproteinase from Vipera ammodytes ammodytes venom.
abstract: A high molecular mass metalloproteinase with α-fibrinogenolytic activity, termed VaF1, was purified from nose-horned viper (Vipera ammodytes ammodytes) venom. Subcutaneous injection of 9 μg of VaF1 did not induce bleeding in rats. Nevertheless, in vitro it degraded collagen IV, nidogen and fibronectin, components of the extracellular matrix, although with low efficacy and narrow specificity. VaF1 would be expected to exert anti-coagulant action, due to its hydrolysis of fibrinogen, factor X, prothrombin and plasminogen, plasma proteins involved in blood coagulation. The enzyme is a single-chain glycoprotein with a molecular mass of 49.7 kDa, as determined by mass spectrometry, and multiple isoelectric points centred at pH 5.8. The complete amino acid sequence of the precursor of VaF1 was deduced by cloning and sequencing its cDNA. Composed of metalloproteinase, disintegrin-like and cysteine-rich domains, VaF1 is a typical P-IIIa subclass snake venom metalloproteinase. Although it possesses a collagen-binding sequence in its disintegrin-like domain, VaF1 displayed no effect on collagen-induced platelet aggregation in vitro. Two consensus N-glycosylation sites are present in the sequence of VaF1, however, the extent of its glycosylation is low, only 5.2% of the total molecular mass. Interestingly, in standard experimental conditions VaF1 is not recognised by antiserum against the whole venom, so it can contribute to post-serotherapy complications, such as ineffective blood coagulation, in the envenomed patient.
authors.affiliation: Department of Molecular and Biomedical Sciences, Jožef Stefan Institute, Jamova cesta 39, SI-1000 Ljubljana, Slovenia.
authors.forename: Adrijana
authors.initials: A
authors.lastname: Leonardi
authors.affiliation: Department of Molecular and Biomedical Sciences, Jožef Stefan Institute, Jamova cesta 39, SI-1000 Ljubljana, Slovenia.
authors.forename: Tamara
authors.initials: T
authors.lastname: Sajevic
authors.affiliation: Department of Molecular and Biomedical Sciences, Jožef Stefan Institute, Jamova cesta 39, SI-1000 Ljubljana, Slovenia.
authors.forename: Zorica
authors.initials: Z
authors.lastname: Latinović
authors.affiliation: Department of Molecular and Biomedical Sciences, Jožef Stefan Institute, Jamova cesta 39, SI-1000 Ljubljana, Slovenia.
authors.forename: Jože
authors.initials: J
authors.lastname: Pungerčar
authors.affiliation: Institute of Immunology, Inc., Rockefellerova 2, HR-10000 Zagreb, Croatia.
authors.forename: Maja Lang
authors.initials: ML
authors.lastname: Balija
authors.affiliation: Medical Centre Ljubljana, Division of Pediatrics, SI-1525 Ljubljana, Slovenia.
authors.forename: Alenka Trampuš
authors.initials: AT
authors.lastname: Bakija
authors.affiliation: Department of Biochemistry and Molecular and Structural Biology, Jamova cesta 39, SI-1000 Ljubljana, Slovenia.
authors.forename: Robert
authors.initials: R
authors.lastname: Vidmar
authors.affiliation: Centre for Research and Knowledge Transfer in Biotechnology, University of Zagreb, Rockefellerova 10, HR-10000 Zagreb, Croatia.
authors.forename: Beata
authors.initials: B
authors.lastname: Halassy
authors.affiliation: Department of Molecular and Biomedical Sciences, Jožef Stefan Institute, Jamova cesta 39, SI-1000 Ljubljana, Slovenia; Department of Chemistry and Biochemistry, Faculty of Chemistry and Chemical Technology, University of Ljubljana, Aškerčeva cesta 5, SI-1000 Ljubljana, Slovenia. Electronic address: igor.krizaj@ijs.si.
authors.forename: Igor
authors.initials: I
authors.lastname: Križaj
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002417:Cattle; D024141:Collagen Type IV / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005170:Factor X / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism; D006868:Hydrolysis; D007106:Immune Sera / Q000276:immunology / Q000378:metabolism; D045726:Metalloproteases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010958:Plasminogen / Q000378:metabolism; D017433:Protein Structure, Secondary; D059748:Proteolysis; D011516:Prothrombin / Q000378:metabolism; D011817:Rabbits; D017386:Sequence Homology, Amino Acid; D014757:Viper Venoms / Q000201:enzymology* / Q000276:immunology; D017819:Viperidae / Q000235:genetics / Q000378:metabolism*
chemical_list: D024141:Collagen Type IV; D005353:Fibronectins; D007106:Immune Sera; D014757:Viper Venoms; D011516:Prothrombin; D005170:Factor X; D005340:Fibrinogen; D010958:Plasminogen; D045726:Metalloproteases
keywords: Metalloproteinase; Serotherapy; Snake venom; Vipera ammodytes ammodytes; α-Fibrinogenase