Disease relevance of PIGK

• We report that GPI transamidase of Trypanosoma brucei (Tb), a causative agent of African sleeping sickness, shares only three components (TbGAA1, TbGPI8, and TbGPI16) with humans and S. cerevisiae but has two other specific components, trypanosomatid transamidase 1 (TTA1) and TTA2 [1].

High impact information on PIGK

• The GPI transamidase mediates GPI anchoring in the endoplasmic reticulum, by replacing a protein's C-terminal GPI attachment signal peptide with a pre-assembled GPI [2].
• GPI8 encodes for an essential 47 kDa type I membrane glycoprotein residing on the luminal side of the ER membrane [3].
• Reconstitution of class K cells with hGPI8 abolishes their accumulation of GPI precursors and restores C-terminal processing of GPI-anchored proteins [4].
• Also, hGPI8 restores the ability of microsomes from the mutant cells to yield an active carbonyl in the presence of a proprotein which is considered to be an intermediate in catalysis by a transamidase [4].
• The lumenal part of PIG-T/GPI16 apparently consists of a beta-propeller with a central hole that regulates the access of substrate protein C termini to the active site of the cysteine protease PIG-K/GPI8 (gating mechanism) as well as of a polypeptide hook that embraces PIG-K/GPI8 [5].

Biological context of PIGK

• GPI transamidase is a key enzyme of this posttranslational modification [6].
• Assignment of phosphatidylinositol glycan, class K (PIGK) gene to porcine chromosome 6q32 by somatic cell and radiation hybrid panel mapping [7].
• GPI8 and PIG-T mutants in which relevant cysteines were replaced with serines were unable to fully restore the surface expression of GPI-anchored proteins upon transfection into their respective mutant cells [8].
• Active site determination of Gpi8p, a caspase-related enzyme required for glycosylphosphatidylinositol anchor addition to proteins [9].

Anatomical context of PIGK

• To definitively characterize this product, RM from K cells transfected with FLAG-tagged GPI8 were employed [10].
• With this system, rough microsomal membranes (RM) containing either [(35)S]-labeled Gaa1p or epitope-tagged Gpi8p, alternative components of the enzymatic complex, were first prepared [10].
• With transamidase competent HeLa cell RM, anti-PLAP or anti-epitope antibody coprecipitated both Gaa1p and Gpi8p consistent with the assembly of the proprotein into a Gaa1p:Gpi8p-containing complex [10].
• GPI transamidase is localized in the endoplasmic reticulum and mediates post-translational transfer of preformed GPI to proteins bearing a carboxyl-terminal GPI attachment signal [8].

Associations of PIGK with chemical compounds

• We also show that cysteine and histidine residues of Gpi8p, which are conserved in members of a cysteine protease family, are essential for generation of a carbonyl intermediate [6].
• We have identified the active site histidine and cysteine residues of leishmanial GPI8 and generated Deltagpi8 lines expressing modified GPI8 proteins [11].
• Ethanolamine phosphate linked to the first mannose residue of glycosylphosphatidylinositol (GPI) lipids is a major feature of the GPI structure that is recognized by human GPI transamidase [12].
• A conserved proline in the last transmembrane segment of Gaa1 is required for glycosylphosphatidylinositol (GPI) recognition by GPI transamidase [13].

Other interactions of PIGK

• When RM from K562 mutant K cells which lack Gpi8p were used, anti-PLAP antibody coprecipitated Gaa1p [10].

Analytical, diagnostic and therapeutic context of PIGK

• Western blots of anti-FLAG bead isolates of solubilized RM from the cells showed that the high Mr band corresponded to Gpi8p covalently bound to miniPLAP [10].

References