J. Biol. Chem., Vol. 266, Issue 18, 11495-11501, 06, 1991
Purification and properties of membrane and cytosolic phosphatidylinositol-specific phospholipases C from human spleen
G Roy, LM Villar, I Lazaro, M Gonzalez, A Bootello and P Gonzalez-Porque
Servicio de Inmunologia, Hospital Ramon y Cajal, Madrid, Spain.
Two phosphatidylinositol-specific phospholipases C (PI-PLC) have been
purified from human spleen. PI-PLCm represents the main activity detected
in the membrane, while PI-PLCc is the main activity present in the
cytoplasm. PI-PLCm can be resolved into two peaks of activity of high Mr
(60,000-70,000) and low Mr (16,000-18,000). High salt concentration
((NH4)2SO4, 2M) dissociates the high Mr form yielding the low molecular
form and increasing the specific activity. The same effect of dissociation
and potentiation of the activity is observed when membranes solubilized by
n-octyl glucoside are subjected to the high voltage conditions of an
isoelectric focusing run. The purified Pi- PLCm has a Mr of about 18,000
when analyzed by sodium dodecyl sulfate- polyacrylamide gel electrophoresis
or gel filtration and a basic pI (9.0-9.2). Purified PI-PLCc has a Mr of
57,000 (sodium dodecyl sulfate- polyacrylamide gel electrophoresis or gel
filtration) and a slightly acid pI (6.2). Other characteristics of both
enzymes, such as cations dependence, substrate specificity, optimum pH, and
kinetic parameters, are also discussed.
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