proteins in supported lipid layers: these proteins, attached to The formation of supported lipid layers incorporating promastithe membrane via lipid moieties, may be less influenced by gote surface protease (PSP) , a glycosylphosphatidylinositol-ansubstrate/protein interactions. chored protein, is investigated using surface plasmon resonance.
A lipid-anchored protein suitable for such studies is pro-Both hydrophilic and hydrophobic substrates are used for the mastigote surface protease (PSP). PSP is the major memformation of lipid layers, and results are consistent with the forbrane protein of the promastigote stage of the protozoan mation of lipid bilayers and monolayers, respectively. Specific parasite Leishmania (11). It is a glycoprotein of 63 kDa antibody binding to layers containing PSP is observed, whereas anchored in the plasma membrane via glycosylphosphatidylnonspecific binding of the antibody to the surface is effectively inositol (GPI) moieties. PSP forms dimers at the plasma suppressed by the phosphatidylcholine lipid layer. Phosphatidylinositol-specific phospholipase C is used to remove the lipid moie-membrane, which can be released into the aqueous phase ties from the membrane-incorporated PSP, releasing it into soluby the action of phosphatidylinositol-specific phospholipase tion in a hydrophilic form and demonstrating that a large fraction C (PIPLC), which removes the lipid moieties from the proof the protein is anchored in the lipid layer via the lipid moieties. tein (12). Antibodies that react with PSP are found in the α§ 1997 Academic Press sera of those with leishmaniasis. Recent studies have demon-Key Words: surface plasmon resonance; supported lipid layers; strated that immunotests based on PSP can be used to distinlipid-anchored membrane proteins; promastigote surface protease. guish between patients with ongoing Leishmania infection and uninfected or cured patients (13, 14).
In this paper we report on investigations of the reconstitution of PSP in supported lipid layers using surface plasmon