The plasma membrane (PPM) and the parasitophorous vacuolar membrane (PVM) of Plasmodium fulfill a number of functions that govern the interactions between host and parasite including nutrient transport, transmembrane signal transduction, and membrane traffic between host and parasite membranes. Given the unique growth requirements of intracellular parasites, these membrane processes may be exploitable in developing novel antimalarial drugs. However, little is known concerning the composition and function of these membranes because of difficulties in their isolation. During the past grant period, an immuno-affinity procedure has been developed to purify the PVM. Using this technique and a variation to isolate the PPM, it is proposed: (1) A general physiological characterization of the PPM and PVM in terms of (1) relative solute permeabilities; (b) identification with specific inhibitors transport and ATPase activities of the PVM/PPM; (c) characterize by metabolic labeling and 2-dimensional electrophoresis PVM and PPM proteins as a function of parasite maturation; and (d) PVM lipid bilayer asymmetry and its maintenance by ATP dependent aminophospholipid translocases. (2) An adenine nucleotide translocator identified in either the PVM or PPM is involved in regulating metabolic cooperation between host cell and parasite metabolism. We propose to determine (a) its localization in either the PVM or PPM; (b) its biosynthesis during asexual maturation and gametocytogenesis; (c) regulation of its transport activity; and (d) its gene structure. (3) Detailed analysis of a PPM electrogenic H+-ATPase in terms of (a) regulation by Ca2+, pH, glucose, and cAMP; (b) activity as a function of parasite maturation; (c) biosynthesis and posttranslational modification as investigated by immunological and molecular probes; and (d) using digitized video-intensified fluorescence microscopy, examine the role of the H+-ATPase in regulating intracellular pH and [Ca2+] during parasite development, glucose deprivation and recovery and gametocyte formation.
