52] Phosphatidylcholine transfer protein from bovine liver

Abstract

Phosphatidylcholine transfer protein (PC-TP) is one of a series of soluble proteins that facilitate the intracellular transfer of lipids between membranes. The PC-TP from bovine liver functions as a specific carder of phosphatidylcholine by forming a 1:1 molecular complex. The protein catalyzes a strict exchange reaction between membranes that contain phosphatidylcholine. Inherent to its mode of action, PC-TP only transfers phosphatii dylcholine that is present in the outer monolayer of membranes. This property has made PC-TP a useful tool in studies on the distribution and transbilayer movement of phosphatidylcholine in membranes. This chapter describes the large-scale purification of PC-TP from bovine liver. The purification of PC-TP is followed by measuring the transfer of [14C]phosphatidylcholine from rat liver microsomes to phospholipid vesicles. PC-TP has also been purified by the use of the vesicle–mitochondrion assay. The phospholipid vesicles containing [32P]phosphatidylcholine and a trace of [14C]triolein as nonexchangeable marker are incubated with beef heart mitochondria as acceptor membrane. After sedimentation of the mitochondria the 32P: 14C ratio of the supernatant is determined. The decrease of this ratio relative to its value before incubation is a measure of PC-TP transfer activity.