The role of inositol acylation and inositol deacylation in GPI biosynthesis in Trypanosoma brucei.
The compound diisopropylfluorophosphate (DFP) selectively inhibits an inositol deacylase activity in living trypanosomes that, together with the previously described phenylmethylsulfonyl fluoride (PMSF)-sensitive inositol acyltransferase, maintains a dynamic equilibrium between the glycosylphosphatidylinositol (GPI) anchor precursor, glycolipid A [NH2(CH2)2PO4-6Man alpha 1-2Man alpha 1-6Man alpha 1-4GlcN alpha 1-6myo-inositol-1-PO4-sn-1,2-dimyristoylglycerol], and its inositol acylated form, glycolipid C. Experiments using DFP in living trypanosomes and a trypanosome cell-free system suggest that earlier GPI intermediates are also in equilibrium between their inositol acylated and nonacylated forms. However, unlike mammalian and yeast cells, bloodstream form trypanosomes do not appear to produce an inositol acylated form of glucosaminylphosphatidylinositol (GlcN-PI). A specific function of inositol acylation in trypanosomes may be to enhance the efficiency of ethanolamine phosphate addition to the Man3GlcN-(acyl)PI intermediate. Inositol deacylation appears to be a prerequisite for fatty acid remodelling of GPI intermediates that leads to the exclusive presence of myristic acid in glycolipid A and, ultimately, in the variant surface glycoprotein (VSG). In the presence of DFP, the de novo synthesis of GPI precursors cannot proceed beyond glycolipid C' (the unremodelled version of glycolipid C) and lyso-glycolipid C'. Under these conditions glycolipid C'-type GPI anchors appear on newly synthesized VSG molecules. However, the efficiencies of both anchor addition to VSG and N-glycosylation of VSG were significantly reduced. A modified model of the GPI biosynthetic pathway in bloodstream form African trypanosomes incorporating these findings is presented.
Güther
M L
ML
Department of Biochemistry, University of Dundee, UK.
Ferguson
M A
MA
D000215:Acylation / Q000187:drug effects; D000818:Animals; D005227:Fatty Acids / Q000378:metabolism; D006017:Glycolipids / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism; D006031:Glycosylation; D017261:Glycosylphosphatidylinositols / Q000096:biosynthesis* / Q000737:chemistry; D007294:Inositol / Q000378:metabolism*; D007531:Isoflurophate / Q000494:pharmacology; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D051381:Rats; D014346:Trypanosoma brucei brucei / Q000187:drug effects / Q000378:metabolism*; D014415:Tunicamycin / Q000494:pharmacology
D005227:Fatty Acids; D006017:Glycolipids; D006023:Glycoproteins; D017261:Glycosylphosphatidylinositols; D014415:Tunicamycin; D007531:Isoflurophate; D007294:Inositol; D010664:Phenylmethylsulfonyl Fluoride