Monday, September 10, 2007

The role of inositol-acylation and inositol-deacylation in the Toxo glycosylphosphatidylinositol biosynthetic pathway

J Biol Chem. 2007 Sep 5; [Epub ahead of print]

The role of inositol-acylation and inositol-deacylation in the Toxoplasma gondii glycosylphosphatidylinositol biosynthetic pathway

Smith TK, Kimmel JF, Azzouz N, Shams-Eldin H, Schwarz RT

College of Life Sciences, University of Dundee, Dundee DD1 5EH.

Toxoplasma gondii is a ubiquitous parasitic protozoan that invades nucleated cells in a process thought to be in part due to several surface, glycosylphosphatidylinositol (GPI)-anchored proteins, like the major surface antigen SAG1 (P30), which dominates the plasma membrane. The serine protease inhibitors phenylmethylsulfonylfluoride and diisopropylfluoride were found to have a profound effect on the T. gondii GPI biosynthetic pathway, leading to the observation and characterization of novel inositol-acylated mannosylated GPI intermediates. This inositol-acylation is acyl-CoA dependent and takes place prior to mannosylation, but uniquely for this class of inositol-acyltransferase it is inhibited by phenylmethylsulfonylfluoride. The subsequent inositol-deacylation of fully mannosylated GPI intermediates is inhibited by both phenylmethylsulfonylfluoride and diisopropylfluoride. The use of these serine protease inhibitors allows observations as to the timing of inositol-acylation and subsequent inositol-deacylation of the GPI intermediates. Inositol-acylation of the non-mannosylated GPI intermediate GlcN-PI precedes mannosylation. Inositol-deacylation of the fully mannosylated GPI intermediate allows further processing, i.e. addition of GalNAc side chain to the first mannose. Characterization of the phosphatidylinositol moieties present on both free GPIs and GPI-anchored proteins shows the presence of a diacyl-glycerol lipid, whose sn-2 position contains almost exclusively an C18:1 acyl chain. The data presented here identifies key novel inositol-acylated mannosylated intermediates, allowing the formulation of an updated T. gondii GPI biosynthetic pathway, along with identification of the putative genes involved.

PMID: 17804418 [PubMed - as supplied by publisher]

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