Phosphoinositide-specific phospholipase C-delta 1: effect of monolayer surface pressure and electrostatic surface potentials on activity

Biochemistry. 1992 Dec 29;31(51):12748-53. doi: 10.1021/bi00166a006.

Abstract

We added phospholipase C-delta 1 (PLC-delta) to the aqueous subphase beneath monolayers formed from mixtures of phosphatidylinositol 4,5-bisphosphate (2% PIP2), phosphatidylserine (33% PS), and phosphatidylcholine (65% PC) and then measured the initial rate of hydrolysis of PIP2 after addition of 10 microM free calcium. Increasing the surface pressure of the monolayer, pi, from 20 to 40 mN/m decreased the rate of hydrolysis 200-fold. The rate of hydrolysis depends exponentially on the surface pressure: rate alpha exp(-pi Ap/kT) where k is the Boltzmann constant, T is the temperature, and Ap congruent to 1 nm2. Similar results were obtained with different (1 and 100 microM) free [Ca2+] and with different mole fractions of PIP2. The results are consistent with a model in which PLC-delta binds to PIP2 with high affinity (Ka = 10(6) M-1) in the absence of calcium ions [Rebecchi, M.J., Peterson, A., & McLaughlin, S. (1993) Biochemistry (preceding paper in this issue)], and a portion of PLC-delta of area Ap inserts into the monolayer doing work = pi Ap prior to hydrolysis of PIP2. Removing the monovalent acidic lipid PS from the monolayer decreases the activity of PLC-delta 4-fold, this effect of PS on activity is similar to the effect of monovalent acidic lipids on the binding of PLC-delta to PIP2 in bilayer vesicles.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Electrochemistry
  • Hydrolysis
  • Kinetics
  • Phosphatidylcholines / metabolism
  • Phosphatidylinositol 4,5-Diphosphate
  • Phosphatidylinositols / metabolism*
  • Phosphatidylserines / metabolism
  • Pressure
  • Surface Properties
  • Type C Phospholipases / chemistry*
  • Type C Phospholipases / metabolism*

Substances

  • Phosphatidylcholines
  • Phosphatidylinositol 4,5-Diphosphate
  • Phosphatidylinositols
  • Phosphatidylserines
  • Type C Phospholipases