Isolation and characterization of two new non-hemorrhagic metalloproteinases with fibrinogenolytic activity from the mapanare (Bothrops colombiensis) venom.
Colombienases are acidic, low molecular weight metalloproteinases (Mr of 23,074.31 Da colombienase-1 and 23,078.80 Da colombienase-2; pI of 6.0 and 6.2, respectively) isolated from Bothrops colombiensis snake venom. The chromatographic profile in RP-HPLC and its partial sequence confirmed its high homogeneity. Both colombienases present fibrino(geno)lytic activity, but did not show any hemorrhagic, amidolytic, plasminogen activator or coagulant activities, and no effect on platelet aggregation induced by collagen or ADP. Both enzymes were strongly active on fibrinogen Aα chains followed by the Bβ chains, and colombienases-2, at high doses, also degraded the γ chains. This activity was stable at temperatures ranging between 4 and 37 °C, with a maximum activity at 25 °C, and at pHs between 7 and 9. The homology demonstrated by the comparison of sequences, with zinc-dependent metalloproteinases, as well as the metal chelant effects on, confirmed that the colombienases were metalloproteinases, particularly to α-fibrinogenases belonging to the P-I class of SVPMs (20-30 kDa), which contain only the single-domain proteins. The biological characteristics of the colombienases confer a therapeutic potential, since they contain a high fibrino(geno)lytic activity, devoid of hemorrhagic activity. These metalloproteinases might be explored as thrombolytic agents given that they dissolve fibrin clots or prevent their formation.
Laboratorio de Inmunoquímica y Ultraestructura, Instituto Anatómico de la Universidad Central de Venezuela, Caracas, Venezuela.
María E
ME
Girón
Alexis
A
Rodríguez-Acosta
Ana María
AM
Salazar
Elda E
EE
Sánchez
Jacob
J
Galán
Carlos
C
Ibarra
Belsy
B
Guerrero
D000595:Amino Acid Sequence; D000818:Animals; D017837:Bothrops; D002851:Chromatography, High Pressure Liquid; D003435:Crotalid Venoms / Q000201:enzymology*; D005340:Fibrinogen / Q000378:metabolism; D005343:Fibrinolytic Agents / Q000494:pharmacology; D006470:Hemorrhage / Q000139:chemically induced; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008297:Male; D045726:Metalloproteases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010974:Platelet Aggregation / Q000187:drug effects; D017386:Sequence Homology, Amino Acid; D013696:Temperature
D003435:Crotalid Venoms; D005343:Fibrinolytic Agents; C464794:fibrinogen Aalpha; D005340:Fibrinogen; D045726:Metalloproteases