M2 (302). GlcNAc-PI is de-N-acetylated to GlcNPI by a deacetylase, PIG-L (step

two) (43). The exact remodeling reaction and also the enzyme that catalyzes the reaction are but to be clarified (see beneath). The Man1 is transferred towards the remodeled GlcN-(acyl) PI from dolichol-phosphate-Man (Dol-P-Man), generating Man-GlcN-(acyl)PI (step six in Fig. two) (44). A complicated of PIG-M, a catalytic element, and PIG-X, a stabilizing component, is GPI mannosyltransferase 1 that catalyzes this reaction (45, 46). The second Man (Man2) is transferred from Dol-P-Man by PIG-V, GPI mannosyltransferase two, producing Man-Man-GlcN-(acyl)PI (step 7 in Fig. two) (47). A side-branch EtNP is attached towards the 2-position ofFig. two. Biosynthesis and protein-attachment of GPI within the ER of mammalian cells. The comprehensive precursor of GPI, termed H8, is synthesized from PI by 11 stepwise reactions and en bloc transferred to proteins. Extra than 20 PIG genes, shown above the biosynthetic pathway, are involved. Modified from Figure 2 in (197) with permission.Journal of Lipid Investigation Volume 57,Man1 to produce Man-(EtNP)Man-GlcN-(acyl)PI (step eight in Fig. 2). The donor of EtNP is PE and PIG-N is GPI-EtNP transferase 1 that catalyzes this step (48). The Man3 is transferred from Dol-P-Man by PIG-B, GPI mannosyltransferase 3, creating Man-Man-(EtNP)Man-GlcN-(acyl)PI (step 9 in Fig. two) (49). The so-called "bridging EtNP" that links GPI to E PIG-W activity (194). In vivo efficacy of this agent has been proteins is attached for the 6-position of Man3 from PE by GPI-EtNP transferase two consisting of PIG-O, a catalytic component, and PIG-F, a stabilizing component (step ten in Fig. 2) (502). The solution, EtNP-Man-Man(EtNP)Man-GlcN-(acyl)PI, is termed H7 and is competent for attachment to proteins (53). H7 is generally further modified by a side-branch EtNP attached towards the 6-position of Man2 to produce EtNP-Man-(EtNP)Man-(EtNP)ManGlcN-(acyl)PI, termed H8 (53). This reaction is mediated by GPI-EtNP transferase 3 consisting of PIG-G, a catalytic element, and PIG-F, a stabilizing element (step 11 in Fig. two) (54). H8 could be the "mature" precursor generally applied to modify proteins. In some GPI-APs, Man4 is attached. Although the precise step at which Man4 is transferred has not been determined in the mammalian system, Man4 in yeast is transferred just before attachment of the bridging EtNP (55).M2 (302). GlcNAc-PI is de-N-acetylated to GlcNPI by a deacetylase, PIG-L (step 2 in Fig. 2) (33, 34). GlcNPI is subsequent translocated for the luminal face by an unknown mechanism (step 3 in Fig. two). It really is believed to be mediated by a "flippase," which has not been identified (35). Lots of of genes involved in GPI biosynthesis had been cloned by expression cloning, taking benefit of mutant Chinese hamster ovary (CHO) cells and other cells defective in each and every among the list of biosynthetic methods (36). Even so, cells defective in the "flipping" step have in no way been established, though the mutant screen seems to become currently saturated (370). It seems that either a putative flippase in addition includes a role essential for cell survival or the flip is mediated by redundant enzymes.