Effect of Monolayer Surface Pressure on the Activities of Phosphoinositide-Specific Phospholipase C-β₁, -γ₁, and -δ₁

Three isoforms of phospholipase C, either PLC-β₁, PLC-γ₁, or PLC-δ₁, were added to the aqueous subphase beneath phospholipid monolayers formed at an air-solution interface, and the initial rate of hydrolysis of phosphatidylinositol 4,5-bisphosphate was measured after addition of 10 µM free Ca²⁺. The monolayers were formed from mixtures of phosphatidylcholine (65% PC), phosphatidylserine (33% PS), and phosphatidylinositol 4,5-bisphosphate (2% PIP₂). Increasing the surface pressure of the monolayer, π, from 15 to 25 mN/m decreases the rate of hydrolysis 16-, 13-, and 5-fold for PLC-β₁, PLC-γ₁, and PLC-δ₁], respectively. The simplest interpretation of these results is that a portion of each of the enzymes of area A_p must insert into the monolayer, doing work πA_p, prior to hydrolysis of PIP₂; binding studies with simple model compounds of known cross-sectional area are consistent with this interpretation. Removing the monovalent acidic lipid PS from the monolayer decreases the initial rates of hydrolysis of PIP₂ about 3-fold for each PLC isoform, which suggests that negative electrostatic surface potentials increase the PLC activity.