A method for the preparation of Tetrahymena thermophila phospholipase A_1 suitable for large-scale production

摘要

A rapid and economical method for the purification of phospholipase A_1 (PLA_1) from the extracellular medium of the ciliate Tetrahymena thermophila is presented. Essentially, the procedure, here designated as purification by selective interaction (PSI), entails the incubation of media containing PLA_1 with liposomes made of soy bean phospholipids. The PLA_1-lipid complexes are precipitated by the addition of CaCl_2 and collected by centrifugation. Elution of the PLA_1 is effected by treating the complexes with 40% dimethylformamide, a reversible inhibitor of this enzyme, which is easily removed by dialysis. In combination with DEAE cellulose ion exchange chromatography, PSI yielded homogeneous PLA_1 preparations with a 14% recovery and a 416-fold increase in specific activity. This procedure, which can be completed within 1 day, may prove useful for the isolation of phospholipases from other sources. This practical method for the purification of a microbial PLA_1 opens the way to large-scale production of these types of enzyme, which are not as yet commercially available.