PIGK (also known as GPI8) is the catalytic subunit of the multi-subunit glycosylphosphatidylinositol-anchor transamidase (GPI-T) complex, an endoplasmic reticulum membrane enzyme that attaches preassembled GPI anchors to the C-terminus of GPI-anchored proteins. During maturation of GPI-anchored proteins in the ER lumen, GPI-T recognizes a diverse C-terminal GPI-attachment signal peptide (lacking a consensus sequence), cleaves it, and forms a new amide bond between the exposed C-terminal residue (omega-site) and the amino group of the bridging ethanolamine-phosphate of the preassembled GPI, i.e. a transamidation reaction. PIGK is a legumain/caspase-like cysteine protease of the peptidase C13 family: its catalytic residues (nucleophile Cys206 and proton donor His164, completed by Asn58) cleave the signal peptide and form a transient acyl(carbonyl)-enzyme thioester intermediate that is then resolved by attack of the GPI ethanolamine amine. PIGK is a single-pass type I ER membrane protein with its catalytic domain in the ER lumen; it assembles with GPAA1, PIGT, PIGS and PIGU into an equimolar heteropentamer, and a disulfide bond to PIGT contributes to full activity. Bi-allelic loss-of-function variants in PIGK cause an autosomal recessive inherited GPI-deficiency disorder (GPIBD), a neurodevelopmental disorder with hypotonia, cerebellar atrophy, and (in most patients) seizures.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0016255 attachment of GPI anchor to protein | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic (IBA) annotation to the core biological process of PIGK - attachment of GPI anchor to protein. This is well supported by direct evidence for human PIGK and is consistent across the GPI8/PIGK orthologue family. Reason: Correct core BP annotation at an appropriate level of specificity. PIGK is the catalytic subunit of the GPI transamidase that replaces the C-terminal GPI-attachment signal peptide with a preassembled GPI anchor. Supporting Evidence: PMID:34576938 recognizes and cleaves the C-terminal GPI attachment signal of |
| GO:0042765 GPI-anchor transamidase complex | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic (IBA) annotation placing PIGK as part of the GPI-anchor transamidase complex. This matches direct human evidence that PIGK assembles with GPAA1, PIGT, PIGS and PIGU into the GPI-T heteropentamer. Reason: Correct core CC annotation; PIGK is a constitutive subunit of the GPI-T complex. Supporting Evidence: PMID:34576938 GPI-TA consists of five subunits: PIGK, GPAA1, PIGT, PIGS, and |
| GO:0003923 GPI-anchor transamidase activity | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic (IBA) annotation to the core molecular function - GPI-anchor transamidase activity. PIGK is the catalytic component that provides this activity within the GPI-T complex. Reason: Correct core MF term. This is the exact term carried by GOA and is the informative molecular function of PIGK, confirmed by direct experimental evidence. Supporting Evidence: PMID:35165458 functions as the catalytic component |
| GO:0003923 GPI-anchor transamidase activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO (IPR028361, GPI_transamidase) electronic mapping to the core molecular function GPI-anchor transamidase activity. Fully consistent with the curated experimental annotations. Reason: Correct and specific IEA mapping to the core MF term; the GPI_transamidase InterPro signature is diagnostic for this activity. Supporting Evidence: PMID:35165458 functions as the catalytic component |
| GO:0005789 endoplasmic reticulum membrane | IEA GO_REF:0000044 | ACCEPT | Summary: Subcellular-location IEA (UniProtKB-SubCell SL-0097) placing PIGK in the endoplasmic reticulum membrane. This is the experimentally established site of GPI-T action and is corroborated by an EXP annotation to the same term. Reason: Correct core CC term; PIGK is an ER membrane protein whose catalytic domain faces the ER lumen where GPI anchoring occurs. Supporting Evidence: PMID:35551457 are catalyzed by an endoplasmic reticulum membrane GPI transamidase complex |
| GO:0006508 proteolysis | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: InterPro2GO mapping from the Peptidase_C13 domain (IPR001096) to proteolysis. PIGK is genuinely a legumain-like cysteine protease that cleaves the C-terminal GPI-attachment signal peptide, so this is biologically accurate, but proteolysis is only one half of the transamidation reaction and is far less informative than GPI-anchor transamidase activity. Reason: Not incorrect - the C13 peptidase activity underlies signal-peptide cleavage - but it captures only the cleavage step and does not describe the integrated transamidation function; retained as non-core context. Supporting Evidence: PMID:10793132 Gpi8p is a catalytic component that cleaves the GPI attachment signal peptide |
| GO:0008233 peptidase activity | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: InterPro2GO mapping from the Peptidase_C13 domain (IPR001096) to the general MF peptidase activity. PIGK is a cysteine protease of the C13/legumain family (MEROPS C13.005), so the domain-based inference is sound, but peptidase activity is a general parent that undersells the specific transamidase activity. Reason: Accurate at the domain level (C13 cysteine protease) but too general to be a core term; the informative MF is GPI-anchor transamidase activity (GO:0003923). Supporting Evidence: file:human/PIGK/PIGK-uniprot.txt Belongs to the peptidase C13 family |
| GO:0016255 attachment of GPI anchor to protein | IEA GO_REF:0000120 | ACCEPT | Summary: Combined-automated-annotation (ARBA/InterPro) electronic annotation to the core BP attachment of GPI anchor to protein. Redundant with the IBA and IDA annotations to the same term and biologically correct. Reason: Correct core BP annotation, consistent with experimental evidence. Supporting Evidence: PMID:34576938 recognizes and cleaves the C-terminal GPI attachment signal of |
| GO:0042765 GPI-anchor transamidase complex | IEA GO_REF:0000120 | ACCEPT | Summary: Combined-automated-annotation (ARBA/InterPro) electronic annotation placing PIGK in the GPI-anchor transamidase complex. Redundant with, and consistent with, the curated IDA/IBA complex annotations. Reason: Correct core CC annotation; PIGK is a bona fide subunit of the GPI-T complex. Supporting Evidence: PMID:34576938 GPI-TA consists of five subunits: PIGK, GPAA1, PIGT, PIGS, and |
| GO:0005515 protein binding | IPI PMID:10793132 Gaa1p and gpi8p are components of a glycosylphosphatidylinos... | MARK AS OVER ANNOTATED | Summary: IntAct protein-protein interaction (PIGK with GPAA1, UniProtKB:O43292). This documents PIGK-GPAA1 association within the GPI-T complex, which is real, but the bare term protein binding is uninformative as a molecular function. Reason: The interaction (PIGK-GPAA1) is genuine and biologically meaningful, but GO:0005515 protein binding is not an informative MF; the interaction is already captured by GPI-anchor transamidase complex membership (GO:0042765). Retained per curation policy rather than removed. Supporting Evidence: PMID:10793132 Gaa1p and Gpi8p are associated with each other |
| GO:0005515 protein binding | IPI PMID:11483512 PIG-S and PIG-T, essential for GPI anchor attachment to prot... | MARK AS OVER ANNOTATED | Summary: IntAct protein-protein interaction annotations (PIGK with GPAA1/PIGT/PIGS). These document GPI-T subunit associations but the bare protein binding term is uninformative. Reason: Genuine intra-complex interactions, but GO:0005515 is uninformative and redundant with the GPI-anchor transamidase complex CC annotation. Retained per policy. Supporting Evidence: PMID:11483512 PIG-S and PIG-T form a protein complex with GAA1 and GPI8 |
| GO:0005515 protein binding | IPI PMID:12802054 Human PIG-U and yeast Cdc91p are the fifth subunit of GPI tr... | MARK AS OVER ANNOTATED | Summary: IntAct protein-protein interaction annotations (PIGK with GPAA1/PIGT). Documents intra-complex associations of the GPI-T subunits. Reason: Real interaction but uninformative bare protein binding term; complex membership is already annotated (GO:0042765). Retained per policy. Supporting Evidence: PMID:12802054 The GPI transamidase complex affinity-purified from cells |
| GO:0005515 protein binding | IPI PMID:28514442 Architecture of the human interactome defines protein commun... | MARK AS OVER ANNOTATED | Summary: High-throughput interactome (BioPlex-type affinity-purification MS) protein binding annotation (PIGK with GPAA1/PIGT). Large-scale interaction screen capturing GPI-T subunit associations. Reason: Uninformative bare protein binding term from a high-throughput interactome; the biologically meaningful content (GPI-T subunit association) is already captured by complex membership. Retained per policy. Supporting Evidence: PMID:11483512 PIG-S and PIG-T form a protein complex with GAA1 and GPI8 |
| GO:0005515 protein binding | IPI PMID:33961781 Dual proteome-scale networks reveal cell-specific remodeling... | MARK AS OVER ANNOTATED | Summary: High-throughput dual-proteome interactome protein binding annotation (PIGK with GPAA1/PIGT). Large-scale interaction screen. Reason: Uninformative bare protein binding term from a high-throughput interactome; redundant with complex membership. Retained per policy. Supporting Evidence: PMID:11483512 PIG-S and PIG-T form a protein complex with GAA1 and GPI8 |
| GO:0005515 protein binding | IPI PMID:40205054 Multimodal cell maps as a foundation for structural and func... | MARK AS OVER ANNOTATED | Summary: High-throughput multimodal cell-map interactome protein binding annotation (PIGK with GPAA1/PIGT). Large-scale interaction/proximity screen. Reason: Uninformative bare protein binding term from a high-throughput interactome; redundant with complex membership. Retained per policy. Supporting Evidence: PMID:11483512 PIG-S and PIG-T form a protein complex with GAA1 and GPI8 |
| GO:0016255 attachment of GPI anchor to protein | TAS Reactome:R-HSA-162791 | ACCEPT | Summary: Reactome traceable annotation for the GPI-T reaction attaching a GPI anchor to uPAR (a representative GPI-anchored substrate), mapped to attachment of GPI anchor to protein. Correctly represents the core BP. Reason: Correct core BP annotation via a specific curated Reactome pathway event. Supporting Evidence: PMID:34576938 recognizes and cleaves the C-terminal GPI attachment signal of |
| GO:0006506 GPI anchor biosynthetic process | IEA GO_REF:0000041 | ACCEPT | Summary: UniPathway-based electronic annotation (UPA00196) to GPI anchor biosynthetic process. GPI-T catalyzes the terminal transamidation step of GPI-anchor biosynthesis, so this parent BP is accurate. Reason: Correct core BP; the transamidation/attachment step is part of GPI-anchor biosynthesis. Consistent with the UniProt PATHWAY (glycosylphosphatidylinositol-anchor biosynthesis). Supporting Evidence: PMID:35551457 are catalyzed by an endoplasmic reticulum membrane GPI transamidase complex |
| GO:0005789 endoplasmic reticulum membrane | NAS PMID:12802054 Human PIG-U and yeast Cdc91p are the fifth subunit of GPI tr... | ACCEPT | Summary: ComplexPortal (CPX-6503) non-traceable statement locating the GPI-T complex, including PIGK, at the ER membrane. Corroborated by an independent EXP annotation to the same term. Reason: Correct core CC; PIGK/GPI-T resides in the ER membrane. Supporting Evidence: PMID:35551457 are catalyzed by an endoplasmic reticulum membrane GPI transamidase complex |
| GO:0016255 attachment of GPI anchor to protein | NAS PMID:12802054 Human PIG-U and yeast Cdc91p are the fifth subunit of GPI tr... | ACCEPT | Summary: ComplexPortal non-traceable statement annotating the GPI-T complex to attachment of GPI anchor to protein. Consistent with abundant direct evidence for PIGK. Reason: Correct core BP annotation. Supporting Evidence: PMID:34576938 recognizes and cleaves the C-terminal GPI attachment signal of |
| GO:0042765 GPI-anchor transamidase complex | IPI PMID:12802054 Human PIG-U and yeast Cdc91p are the fifth subunit of GPI tr... | ACCEPT | Summary: ComplexPortal-curated complex membership of PIGK in the GPI-anchor transamidase complex (CPX-6503). PMID:12802054 established PIG-U as the fifth subunit and confirmed the affinity-purified GPI8-containing complex. Reason: Correct core CC annotation of PIGK as a subunit of the GPI-T complex. Supporting Evidence: PMID:12802054 The GPI transamidase complex affinity-purified from cells |
| GO:0005789 endoplasmic reticulum membrane | EXP PMID:11483512 PIG-S and PIG-T, essential for GPI anchor attachment to prot... | ACCEPT | Summary: Experimental evidence localizing PIGK/GPI8 (and the GPI-T complex) to the endoplasmic reticulum membrane. This is the definitive experimental support for the ER-membrane location. Reason: Correct core CC term, directly supported by experiment. Supporting Evidence: PMID:11483512 The GPI transamidase mediates GPI anchoring |
| GO:0003923 GPI-anchor transamidase activity | IDA PMID:35551457 Molecular insights into biogenesis of glycosylphosphatidylin... | ACCEPT | Summary: Direct experimental evidence (cryo-EM structure plus structure-based mutagenesis of the reconstituted human GPI-T) for GPI-anchor transamidase activity, with PIGK as the catalytic subunit. This is the primary core MF annotation. Reason: Correct core MF term with strong direct experimental support; matches the exact GOA term. Supporting Evidence: PMID:35551457 suggests a legumain-like |
| GO:0180046 GPI anchored protein biosynthesis | IDA PMID:35165458 Structure of human glycosylphosphatidylinositol transamidase... | ACCEPT | Summary: Direct experimental evidence (structure and functional analysis of human GPI-T) that PIGK participates in GPI anchored protein biosynthesis. GO:0180046 is the current specific BP for the GPI-AP maturation process. Reason: Correct core BP; PIGK is essential for GPI-anchored protein biosynthesis (maturation), as shown by the catalytic dyad requirement. Supporting Evidence: PMID:35165458 which is essential for |
| GO:0180046 GPI anchored protein biosynthesis | IDA PMID:35551457 Molecular insights into biogenesis of glycosylphosphatidylin... | ACCEPT | Summary: Direct experimental evidence that PIGK/GPI-T is required for GPI anchored protein biosynthesis, from the cryo-EM structure and functional mutagenesis of the human complex. Reason: Correct core BP annotation, redundant with and consistent with the other IDA to GO:0180046. Supporting Evidence: PMID:35551457 important step towards the mechanistic understanding of |
| GO:0003923 GPI-anchor transamidase activity | IDA PMID:37684232 Structures of liganded glycosylphosphatidylinositol transami... | ACCEPT | Summary: Direct experimental evidence for GPI-anchor transamidase activity from substrate- and product-bound cryo-EM structures of human GPI-T, defining a caspase-like catalytic mechanism with PIGK as the catalytic subunit (Cys206 nucleophile / His164 proton donor). Reason: Correct core MF term with strong structural/mechanistic support. Supporting Evidence: PMID:37684232 inform a caspase-like catalytic mechanism |
| GO:0016255 attachment of GPI anchor to protein | IDA PMID:12802054 Human PIG-U and yeast Cdc91p are the fifth subunit of GPI tr... | ACCEPT | Summary: Direct experimental evidence that PIGK/GPI-T is required for attachment of GPI anchor to protein. Class-U (PIG-U-deficient) cells lacking a functional GPI-T could not cleave the GPI attachment signal peptide. Reason: Correct core BP annotation supported by loss-of-function cell evidence. Supporting Evidence: PMID:12802054 had no ability to cleave the GPI attachment signal peptide |
| GO:0016255 attachment of GPI anchor to protein | IDA PMID:34576938 Functional Analysis of the GPI Transamidase Complex by Scree... | ACCEPT | Summary: Direct experimental evidence (subunit mutagenesis affecting GPI-anchor attachment to protein) that PIGK is required for attachment of GPI anchor to protein. Reason: Correct core BP annotation; mutagenesis of PIGK catalytic residues (e.g. His164, Cys206) abolishes GPI-anchor attachment. Supporting Evidence: PMID:34576938 recognizes and cleaves the C-terminal GPI attachment signal of |
| GO:0016255 attachment of GPI anchor to protein | IDA PMID:37684232 Structures of liganded glycosylphosphatidylinositol transami... | ACCEPT | Summary: Direct experimental evidence from substrate/product-bound GPI-T structures that PIGK mediates attachment of GPI anchor to protein via the transamidation reaction. Reason: Correct core BP annotation with mechanistic structural support. Supporting Evidence: PMID:37684232 replaces it with GPI via a transamida |
| GO:0042765 GPI-anchor transamidase complex | IDA PMID:37684232 Structures of liganded glycosylphosphatidylinositol transami... | ACCEPT | Summary: Direct experimental (cryo-EM) evidence identifying PIGK as a subunit of the GPI-anchor transamidase complex, resolved together with GPAA1, PIGS, PIGT and PIGU. Reason: Correct core CC annotation with direct structural support. Supporting Evidence: PMID:37684232 The transmembrane complex GPI-T |
| GO:0016255 attachment of GPI anchor to protein | IDA PMID:35165458 Structure of human glycosylphosphatidylinositol transamidase... | ACCEPT | Summary: Direct experimental evidence (human GPI-T structure and mutagenesis of the C206-H164-N58 catalytic triad) that PIGK mediates attachment of GPI anchor to protein. Reason: Correct core BP annotation with strong structural/functional support. Supporting Evidence: PMID:35165458 Attaching GPI to the protein in the endoplasmic reticulum (ER) is catalyzed by |
| GO:0016255 attachment of GPI anchor to protein | IDA PMID:35551457 Molecular insights into biogenesis of glycosylphosphatidylin... | ACCEPT | Summary: Direct experimental evidence (2.53-A cryo-EM structure and structure-based mutagenesis) that PIGK/GPI-T mediates attachment of GPI anchor to protein. Reason: Correct core BP annotation, redundant with and consistent with other IDAs. Supporting Evidence: PMID:35551457 covalent attachment of GPI at the new carboxyl terminus |
| GO:0042765 GPI-anchor transamidase complex | IDA PMID:35165458 Structure of human glycosylphosphatidylinositol transamidase... | ACCEPT | Summary: Direct experimental (cryo-EM) identification of PIGK as the catalytic subunit of the GPI-anchor transamidase complex, resolved with the four other subunits. Reason: Correct core CC annotation with direct structural support. Supporting Evidence: PMID:35165458 The PIGK subunit |
| GO:0042765 GPI-anchor transamidase complex | IDA PMID:35551457 Molecular insights into biogenesis of glycosylphosphatidylin... | ACCEPT | Summary: Direct experimental (cryo-EM) evidence resolving PIGK within the equimolar heteropentameric GPI-anchor transamidase complex. Reason: Correct core CC annotation with direct structural support. Supporting Evidence: PMID:35551457 revealing an equimolar |
| GO:0016255 attachment of GPI anchor to protein | IDA PMID:10793132 Gaa1p and gpi8p are components of a glycosylphosphatidylinos... | ACCEPT | Summary: Direct experimental evidence that GPI8/PIGK is required for GPI attachment; conserved cysteine and histidine residues essential for the carbonyl intermediate identify PIGK as the catalytic component that cleaves the GPI signal peptide. Reason: Correct core BP annotation, supported by mutagenesis of catalytic residues. Supporting Evidence: PMID:10793132 Gpi8p is a catalytic component that cleaves the GPI attachment signal peptide |
| GO:0016255 attachment of GPI anchor to protein | IDA PMID:9356492 The affected gene underlying the class K glycosylphosphatidy... | ACCEPT | Summary: Direct experimental evidence identifying hGPI8/PIGK as the gene defective in the class-K GPI-deficient mutant; reconstitution with hGPI8 restored C-terminal processing of GPI-anchored proteins. Reason: Foundational functional evidence for the core BP; PIGK loss abolishes, and its restoration rescues, GPI anchor attachment. Supporting Evidence: PMID:9356492 restores the ability |
| GO:0042765 GPI-anchor transamidase complex | IDA PMID:12582175 Two subunits of glycosylphosphatidylinositol transamidase, G... | ACCEPT | Summary: Direct experimental evidence that GPI8/PIGK is a subunit of the multimeric mammalian GPI transamidase and forms a functionally important disulfide bond with PIG-T within the complex. Reason: Correct core CC annotation; PIGK-PIGT disulfide within the GPI-T complex. Supporting Evidence: PMID:12582175 is a multimeric complex consisting of at least five subunits |
| GO:0042765 GPI-anchor transamidase complex | IDA PMID:34576938 Functional Analysis of the GPI Transamidase Complex by Scree... | ACCEPT | Summary: Direct experimental evidence (purification and functional analysis of the five-subunit human GPI-TA) confirming PIGK as a subunit of the GPI-anchor transamidase complex. Reason: Correct core CC annotation. Supporting Evidence: PMID:34576938 GPI-TA consists of five subunits: PIGK, GPAA1, PIGT, PIGS, and |
| GO:0042765 GPI-anchor transamidase complex | IDA PMID:12802054 Human PIG-U and yeast Cdc91p are the fifth subunit of GPI tr... | ACCEPT | Summary: Direct experimental evidence that the GPI8/PIGK-containing GPI transamidase complex affinity-purified from cells contains PIG-U and four other components, confirming PIGK complex membership. Reason: Correct core CC annotation. Supporting Evidence: PMID:12802054 The GPI transamidase complex affinity-purified from cells |
| GO:0003923 GPI-anchor transamidase activity | TAS Reactome:R-HSA-162836 | ACCEPT | Summary: Reactome traceable annotation for the GPI-T-catalyzed reaction (uPAR precursor + acyl-GPI -> uPAR-acyl-GPI + propeptide), mapped to GPI-anchor transamidase activity - the exact GOA core MF term. Reason: Correct core MF annotation via a curated Reactome reaction. Supporting Evidence: PMID:35165458 functions as the catalytic component |
| GO:0042765 GPI-anchor transamidase complex | IDA PMID:11483512 PIG-S and PIG-T, essential for GPI anchor attachment to prot... | ACCEPT | Summary: Direct experimental evidence that PIG-S and PIG-T form a complex with GAA1 and GPI8/PIGK, establishing PIGK membership in the GPI-anchor transamidase complex. Reason: Correct core CC annotation. Supporting Evidence: PMID:11483512 PIG-S and PIG-T form a protein complex with GAA1 and GPI8 |
| GO:0016020 membrane | HDA PMID:19946888 Defining the membrane proteome of NK cells. | MARK AS OVER ANNOTATED | Summary: High-throughput mass-spectrometry detection of PIGK in an NK-cell membrane proteome. Consistent with PIGK being membrane-associated but the generic term membrane is far less informative than the established ER membrane location. Reason: Not incorrect (PIGK is a membrane protein) but GO:0016020 membrane is uninformatively general and is superseded by the specific ER membrane (GO:0005789) annotations. Supporting Evidence: PMID:19946888 define the composition of the membrane |
| GO:0005789 endoplasmic reticulum membrane | TAS Reactome:R-HSA-162836 | ACCEPT | Summary: Reactome traceable annotation placing the GPI transamidase (including PIGK) at the ER membrane, consistent with the experimental EXP annotation. Reason: Correct core CC annotation. Supporting Evidence: PMID:35551457 are catalyzed by an endoplasmic reticulum membrane GPI transamidase complex |
| GO:0034235 GPI anchor binding | TAS PMID:10793132 Gaa1p and gpi8p are components of a glycosylphosphatidylinos... | KEEP AS NON CORE | Summary: Traceable annotation that PIGK contributes_to GPI anchor binding. The GPI-T complex binds the preassembled GPI lipid substrate (a composite GPI-binding cavity is resolved in structures), and PIGK contributes to this binding as the catalytic subunit acting on the GPI ethanolamine. Reason: Biologically reasonable with the contributes_to qualifier: GPI binding is a complex-level property to which PIGK contributes, not an independent PIGK activity. Retained as non-core supporting MF. Supporting Evidence: PMID:35551457 an endogenous GPI in the structure defines a composite cavity for the lipid |
| GO:0016255 attachment of GPI anchor to protein | TAS PMID:11483512 PIG-S and PIG-T, essential for GPI anchor attachment to prot... | ACCEPT | Summary: Traceable annotation that PIGK/GPI-T mediates GPI anchor attachment to proteins in the ER by replacing the C-terminal GPI-attachment signal peptide. Reason: Correct core BP annotation. Supporting Evidence: PMID:11483512 The GPI transamidase mediates GPI anchoring |
| GO:0003923 GPI-anchor transamidase activity | IMP PMID:10793132 Gaa1p and gpi8p are components of a glycosylphosphatidylinos... | ACCEPT | Summary: Mutational evidence that conserved catalytic cysteine/histidine residues of GPI8/PIGK are essential for the carbonyl intermediate, i.e. for GPI-anchor transamidase activity. Catalytic-residue mutants abolish activity. Reason: Correct core MF term with genetic/mutational support (loss of transamidase activity on mutating catalytic residues). Supporting Evidence: PMID:10793132 essential for generation of a carbonyl intermediate |
| GO:0042765 GPI-anchor transamidase complex | IMP PMID:10793132 Gaa1p and gpi8p are components of a glycosylphosphatidylinos... | ACCEPT | Summary: Evidence that Gaa1p and Gpi8p/PIGK associate as components of the GPI transamidase, placing PIGK in the GPI-anchor transamidase complex. Reason: Correct core CC annotation. Supporting Evidence: PMID:10793132 Gaa1p and Gpi8p are associated with each other |
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Download this section (compressed HTML)Q: PIGK carries the catalytic triad, yet loss of any one of the other four subunits abolishes activity and the GPI lipid-binding cleft is formed by partner transmembrane helices. Should PIGK's GPI-anchor transamidase activity (GO:0003923) be annotated as enables, or is contributes_to a more faithful model of an activity that only exists in the assembled heteropentamer?
Suggested experts: Kinoshita T, Ohishi K, Hong Y
Q: PIGK is annotated to peptidase activity (GO:0008233) and proteolysis (GO:0006508) by InterPro2GO on the basis of its peptidase C13 fold. Given that the cleavage step is an obligate half-reaction of transamidation and no free proteolytic product accumulates, are these annotations informative or should they be treated as fold-derived over-annotations?
Suggested experts: Kinoshita T, Fujita M, Murakami Y
Q: The RNF121 E3 ligase is resolved bound to the transamidase in one cryo-EM structure and proposed as a quality-control factor. Is RNF121 a bona fide constitutive component of the PIGK complex, and does it ubiquitinate PIGK or a partner subunit?
Suggested experts: Kinoshita T, Zhang H
Q: How does the autoinhibitory loop that occludes the PIGK catalytic site in the resting enzyme relate to the disease-causing PIGK variants in neurodevelopmental disorder with hypotonia and cerebellar atrophy - do pathogenic substitutions destabilize the complex, the triad, or the activation rearrangement?
Suggested experts: Kinoshita T, Murakami Y
Experiment: Express and purify PIGK alone, PIGK-GPAA1, and the full pentamer into nanodiscs and assay transamidation of a defined proprotein substrate with an acyl-GPI donor by mass spectrometry, quantifying both the acyl-enzyme intermediate and the finished GPI-anchored product. If PIGK alone forms no acyl-enzyme intermediate on a physiological substrate, the catalytic step is complex-dependent throughout and not merely lipid-binding-dependent.
Hypothesis: The GPI-anchor transamidase activity is a property of the assembled pentamer rather than of PIGK alone, so PIGK's molecular function is better modelled as contributes_to.
Type: stepwise in vitro reconstitution with intermediate trapping
Experiment: Incubate purified PIGK-containing GPI-T with a proprotein substrate in the presence and absence of the GPI donor, then quantify released C-terminal signal peptide by targeted mass spectrometry. Test whether cleavage occurs at an appreciable rate without an acceptor nucleophile, and repeat with small nucleophiles (hydrazine, hydroxylamine) as a positive control for acyl-enzyme turnover.
Hypothesis: PIGK has no independent proteolytic activity outside transamidation, so the InterPro2GO-derived peptidase and proteolysis annotations describe a fold rather than a function.
Type: donor-dependence proteolysis assay
Experiment: Reconstitute a panel of reported PIGK patient variants in PIGK-null cells and measure in parallel (i) incorporation into the pentamer by blue-native PAGE and co-immunoprecipitation, (ii) surface GPI-anchored protein levels by FLAER and CD59/CD55 flow cytometry, and (iii) in vitro transamidase activity of purified complexes. Map each variant onto the liganded structures to test whether the mechanistic class predicts the biochemical phenotype.
Hypothesis: Pathogenic PIGK variants act through distinct mechanisms - complex destabilization, triad disruption, or failure of the autoinhibitory-loop rearrangement - which would predict different residual activity and different responsiveness to substrate load.
Type: variant panel with complex-assembly, cell-surface and in vitro activity readouts
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