PHOSPHOLIPASE-C ASSOCIATED WITH PARTICULATE FRACTIONS OF BOVINE BRAIN

Abstract

We previously reported that cytosolic fractions of bovine brain contain two immunologically distinct phosphoinositide-specific phospholipases C (PLCs), PLC-I and PLC-II. In this report the subcellular distribution of PLC-I and PLC-II in brain homogenates was measured using RIA. Significant differences were found in the distribution of the two forms of PLC in 100,000 X g supernatants (cytosolic fraction) of brain homogenized in hypotonic buffer and 2 M KCl extracts of washed pellets (particulate fraction). More than 90% of PLC-II was found in the cytosolic fractions, whereas the PLC-I-like molecules were equally distributed between cytosolic and particulate fractions. Purification of PLC enzyme to near homogeneity from the particulate fractions yielded two PLC enzymes, both of which could be recognized by anti-PLC-I antibodies but not by anti-PLC-II antibodies. Their Mr values, determined under denaturing conditions, were 150,000 and 140,000. The polypeptide of the enzyme of Mr 150,000 seems to be the same as that of the cytosolic enzyme PLC-I: their Mr values were identical, and their trypsin-digested peptides yielded a similar elution profile on a C18 reverse-phase column. We propose, therefore, that PLC-I and its truncated form are weakly associated with membranes.