Production & purification of a fibrinolytic enzyme from Bionectria sp.

ROVATI, JI; ARNAU, VG; FIGUEROA, LIC AND FARIÑA, JI

San Miguel de Tucumán

Congreso; XLV Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2009

Several thrombolytic agents such as urokinase, alteplase (t-PA), and streptokinase (SK) are currently available for clinical usage, although they still suffer significant shortcomings, including requeriment of large therapeutic dose, short plasma half-life, limited fibrin specificity, reocclusion and bleeding complications. Among microbial sources, members of the genre Bacillus, Aspergillus, Fusarium and Streptomyces have been reported as fibrinolytic enzyme producers. In the present study we attempted the isolation of a novel fibrinolytic enzyme from cultures of Bionectria sp. LY 4.1, a filamentous fungus from Las Yungas Pedemontana forests. After cultivation under submerged culture conditions at shake-flask scale, supernatant was fractionally precipitated with cold acetone, followed by further purification by gel filtration chromatography with Sephacryl S-300 HR. Fibrinolytic activity was monitored by the fibrin plate method. Quantification of the fibrinolytic activity was determined according to the amount of tyrosine released from fibrin clots per min. The effect of pH and incubation temperature was studied. Finally, in vitro activity over human blood clots was also analyzed to verify thrombolytic effects. Results confirmed that the purified fibrinolytic enzyme obtained from cultures of Bionectria sp LY 4.1 showed thrombolytic effect over human blood clots and constitude a promising candidate for treating thrombolytic diseases with the additional benefits of its more specific mode of action (plasminogen independent) and probably associated to less side effects.