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ENZYMATIC HYDROLYSIS OF IMMOBILIZED SPHINGOMYELIN BY THREE BACTERIAL PHOSPHOLIPASES C
Author(s) -
MALMQVIST TORSTEN,
MALMQVIST MAGNUS,
MÖLLBY ROLAND
Publication year - 1981
Publication title -
acta pathologica microbiologica scandinavica section b microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.909
H-Index - 88
eISSN - 1600-0463
pISSN - 0304-131X
DOI - 10.1111/j.1699-0463.1981.tb00200_89b.x
Subject(s) - sphingomyelin , enzyme , chemistry , hydrolysis , biochemistry , phospholipase , enzymatic hydrolysis , chromatography , microbiology and biotechnology , biology , membrane
Through hydrophobic interaction, sphingomyelin was adsorbed to agarose beads containing octyl groups by a stepwise dilution procedure. This immobilized lipid was used as a substrate for three bacterial phospholipases C (E.C. 3. 1. 4. 3.). The degradation with time of this substrate showed two different fractions of the substrate according to hydrolysing velocity in the early part of the time‐curve when phospholipases C from Bacillus cereus and Clostridium perfringens were used. The early fractions could be predigested by the enzymes, a procedure which resulted in linear time‐curves. The corresponding early part of the time‐curve for phospholipase C from Staphylococcus aureus was linear, indicating a comparatively large early fraction of the substrate for this enzyme. The stock gel of the immobilized lipid substrate could be stored for months. It was easily and reproducibly handled as a water suspension. After enzymatic hydrolysis the substrate was rapidly separated from enzyme and product by filtration. The enzyme assay presented thus represents a convenient way to avoid the difficulties connected with the use of temporary sonicated suspensions as substrate for bacterial phospholipases C.