Phospholipid-exchange proteins for the topological distribution of microsomal phospholipids [proceedings].

The immunological method was therefore shown to be capable of isolating and partially purifying membranes from a whole rat liver with approx. 40% yield within 50min from breaking cells and with only minimal exposure to temperatures below physiological ones. Electron microscopy of the membrane preparation showed organelles, including mitochondria, sometimes enclosed inside larger membranous elements. This may partly explain the apparently specific binding of non-plasma-membrane elements to the immunoadsorbent, and resulting impurity of the plasmamembrane preparation. The membrane isolated on immunoadsorbent retained sensitivity to glucagon, as shown by measurement of adenylate cyclase activity (Table 2). The specific activity of this enzyme and degree of stimulation by fluoride and glucagon was comparable with values obtained for liver plasma membranes purified by other techniques (Pohl et al., 1969). The recovery of cyclic AMP phosphodiesterase, assayed at a substrate concentration of 1 C(M, was very low in the immunoadsorbent-membrane pellet relative to recoveries of plasma-membrane markers (Table 1). However, in other experiments it was shown that approx. 50 % of the total liver-cell-homogenate phosphodiesterase, measured with 1 pM-CyCliC AMP, was recovered in the pellet obtained after centrifugation for 90min at 1OOOOOg. This suggests that membrane-bound cyclic AMP phosphodiesterase in rat liver is associated primarily with intracellular membranes rather than the plasma membrane, as also proposed for the enzyme in fat-cells (Kono et al., 1975). The fraction of phosphodiesterase active at low ( PM) substrate concentrations that is particulate appears to be considerably less in liver than in adipose tissue.