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SOLUBILIZATION AND CHARACTERIZATION OF MICROSOMAL‐ASSOCIATED PHOSPHATIDYLINOSITOL: CERAMIDE PHOSPHOINOSITOL TRANSFERASE FROM SACCHAROMYCES CEREVISIAE
Author(s) -
KO JESANG,
CHEAH SHWUYENG,
FISCHL ANTHONY S.
Publication year - 1995
Publication title -
journal of food biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.507
H-Index - 47
eISSN - 1745-4514
pISSN - 0145-8884
DOI - 10.1111/j.1745-4514.1995.tb00533.x
Subject(s) - ceramide , phosphatidylinositol , biochemistry , sphingolipid , chemistry , enzyme , yeast , transferase , microsome , saccharomyces cerevisiae , enzyme assay , kinase , apoptosis
The membrane associated enzyme phosphatidylinositol: ceramide phosphoinositol transferase (IPC synthase) catalyzes an essential step in the biosynthesis of yeast inositol‐containing sphingolipids. A variety of solubilization agents were examined for their ability to release IPC synthase activity from yeast membranes. The most effective solubilization agent was Triton X‐100 which released over 90% of the IPC synthase activity. The basic enzymological properties of the solubilized enzyme were determined using a Triton X‐100/phosphatidylinositol/ceramide/enzyme mixed micellar assay system. IPC synthase activity was dependent upon the surface concentrations of phosphatidylinositol and ceramide in a phosphatidylinositol/ceramide/Triton X‐100 mixed micelle. Maximal enzyme activity was measured at 30C and pH 7.0 in the presence of 5 mM Triton X‐100, 1 mM manganese and 5 mM magnesium ions.