An efficient method for purification of lumbrokinase from the earthworm Eisenia foetida

IANNUCCI, NANCY BEATRIZ; CAMPERI, SILVIA ANDREA; CASCONE, OSVALDO

Rosario

Congreso; XLII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2006

SAIB

  Fibrinolytic agents are widely used for thrombosis prevention and treatment. Lumbrokinases (LK) are thrombolytic enzymes from earthworms suitable for oral administration due to their high stability in the gastrointestinal tract. In this work we report a novel purification method for LK from the earthworm Eisenia foetida. Earthworms were homogenised with water. Polyethyleneglycol (PEG), monosodium phosphate and dipotassium phosphate were added to build an aqueous twophase system. The clear top phase, enriched in PEG, contained all the LK activity together with other proteins and the bottom phase contained the homogenised tissue and the earth. The upper phase was loaded on a Q-Sepharose column equilibrated with 20 mM sodium phosphate buffer, pH 5.8. LK elution was achieved by addition of NaCl up to 0.5 M to the equilibrating buffer. The fraction containing LK was loaded to a Benzamidine- Sepharose column equilibrated with the elution buffer of the previous column. The LK was eluted with 0.1 M sodium acetate. The yield was 90 % and the purification factor 9. SDS-PAGE showed a single band corresponding to the LK molecular weight.