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Abstract
Glycosylphosphatidylinositol (GPI) cleavage by a GPI phospholipase C (GPIPLCp) is suppressed in Trypanosoma brucei. However, heterologous expression of GPIPLCp in Leishmania results in a GPI deficiency, indicating an absence of regulation of the phospholipase activity against GPIs.|The purpose of this study was to determine possible mechanisms governing regulation of GPI-PLCp activity against GPIs in vivo using Leishmania as a model system. We identified several criteria that affect the ability of GPI-PLCp to cleave GPIs in vivo: i) Cys80 or ii) at least two Cys within a C-terminal cluster (positions 269,270, 273) must be present. A specific intracellular location and activity of GPI-PLCp were needed for the enzyme to cause the GPI-negative phenotype.|GPI-PLCp is an integral membrane protein. The affinity of the enzyme for biological membranes allows it to gain access to GPIs. Therefore, it is important to understand the mechanism by which GPI-PLCp binds to membranes. We report the presence of two regions capable of post-translationally targeting a soluble reporter to T. brucei membranes. We propose a model in which GPI-PLCp is anchored in the membrane by "hydrophobic patches" created by the amphipathic helices in the protein.