Stabilization of quaternary structure and activity of bovine liver catalase through encapsulation in liposomes

Abstract

Bovine liver catalase was encapsulated in an aqueous phase of the phospholipid vesicle (liposome) to improve the stability of its tetrameric structure and activity. The catalase-containing liposomes (CALs) prepared were 30, 50 and 100 nm in mean diameters (CAL 30, CAL 50 and CAL 100, respectively). The CAL 100 included the types I, II and III based on the amounts of catalase encapsulated. The CAL 30, CAL 50 and CAL 100-I contained one catalase molecule per liposome, and the CAL 100-II and CAL 100-III on average 5.2 and 17 molecules, respectively. The storage stability of catalase in either CAL system was significantly increased compared to that of free catalase at 4 °C in a buffer of pH 7.4. At 55 °C, free catalase was much more deactivated especially with decreasing its concentration predominantly due to enhanced dissociation of catalase into subunits while it was so done at excessively high enzyme concentration mainly due to enhanced formation of catalase intermolecular aggregates. Among the three types of CAL 100, the CAL 100-II showed the highest thermal stability, indicating that an excess amount of catalase in the CAL 100-III was also disadvantageous to maintain an active form of the catalase even in liposome. In the CAL 100-III, however, the stability of catalase was significantly improved compared to that of free catalase at the same concentration. The CAL thermal stability was little affected by the liposome size as observed in the CAL 30, CAL 50 and CAL 100-I. An intrinsic tryptophan fluorescence of the catalase recovered from the CAL 100-II thermally treated at 55 °C revealed that a partially denatured catalase molecule was stabilized through its hydrophobic interaction with liposome membrane. This interaction depressed not only dissociation of catalase into subunits but also formation of an inactive intermolecular aggregate between the catalase molecules in a liposome. Furthermore, either type of CAL 100 showed a higher stability than free catalase in the successive decompositions of 10 mM H 2O 2 at 25 °C mainly because the H 2O 2 concentration was kept low inside liposomes due to the permeation barrier of the lipid membrane to H 2O 2.