EOGT

Summary: The inherited ER-lumen localization agrees with experimentally tested fly Eogt and the targeting features of canonical human EOGT.

Reason: The two EOGT IBA assertions reside at the same PAINT node, PTN001841612. The current PANTHER 19 tree places human Q5NDL2 below this node, and the ungapped target MSA sequence exactly matches the 527-residue canonical protein; the pinned August 2026 IBD contains the expected FlyBase donor and no matching NOT assertion on the target path. In PMID:22158438, Results/Figure 3c–f and Methods, fly Eogt in S2 cells colocalizes with GFP-KDEL, while deletion of its terminal RNEL or C-terminal tagging shifts the protein into culture medium. These are fly assays, not direct human microscopy. Human Q5NDL2-1 has an N-terminal signal sequence and terminal HDEL consistent with the conserved location. The activity and location of shorter natural products are not established by this inference. Independent mouse experiments in PMID:25488668 show expressed EOGT in the ER (Results/Figure 1B) and the O-GlcNAc product signal overlapping calnexin (Figure 1C); purified-enzyme transfer is measured in Figure 2. The expressed enzyme is mouse even when HEK293T cells are the host.

Summary: The UniProt subcellular-location mapping correctly specifies the ER lumen for the canonical EOGT enzyme.

Reason: Current SL-0096 maps directly to GO:0005788. The location is consistent with the canonical human signal sequence and HDEL retrieval motif, the conserved PAINT placement, and the fly localization and secretion experiments in PMID:22158438 Figure 3c–f. The mature human chain is annotated at 18–527 by sequence analysis; its cleavage site is not independently measured here. The mapping should not be read as experimentally establishing the localization of the 208-residue product, which lacks the canonical C-terminal retrieval motif. PMID:25488668 independently shows mouse EOGT in the ER (Results/Figure 1B), the glycosylation-product signal overlapping calnexin (Figure 1C), and purified-enzyme catalysis (Figure 2). These are mouse ortholog experiments, including the HEK293T-hosted microscopy.

Summary: Orthology-based protein O-linked glycosylation is supported by donor provenance and direct human EOGT assays.

Reason: EOGT catalyzes the initiating transfer itself, rather than merely being a glycosylated substrate. The official mouse donor Q8BYW9 maps to MGI:2141669 and Ensembl ENSMUSP00000061610.5 and carries direct IDA annotations to this process and GO:0097363 from PMID:22310717. That older paper is abstract-only in the normal cache; its full Methods were not recovered, so its assay detail is not claimed. Independently read PMID:23671640 Results/Figure 1 and Methods demonstrate that the human 527-residue clone restores O-GlcNAc modification of Notch EGF repeats in fly S2 cells, establishing the same catalytic process on the human product. Retain GO:0006493: the current narrower GO:7770074 explicitly restricts the product to an unextended single sugar, whereas the mammalian donor report describes further galactosylation. This is a definition-based specificity limit, not evidence that EOGT itself performs the downstream galactosylation. Full Methods and Results in PMID:25488668 additionally establish catalysis by purified CHO-expressed mouse EOGT using radiolabeled UDP-GlcNAc and a fly Notch EGF20 substrate, providing a separately read primary mouse assay.

Summary: The manually transferred O-linked glycosylation process is established for human EOGT.

Reason: EOGT catalyzes the initiating transfer itself, rather than merely being a glycosylated substrate. The official mouse donor Q8BYW9 maps to MGI:2141669 and Ensembl ENSMUSP00000061610.5 and carries direct IDA annotations to this process and GO:0097363 from PMID:22310717. That older paper is abstract-only in the normal cache; its full Methods were not recovered, so its assay detail is not claimed. Independently read PMID:23671640 Results/Figure 1 and Methods demonstrate that the human 527-residue clone restores O-GlcNAc modification of Notch EGF repeats in fly S2 cells, establishing the same catalytic process on the human product. Retain GO:0006493: the current narrower GO:7770074 explicitly restricts the product to an unextended single sugar, whereas the mammalian donor report describes further galactosylation. This is a definition-based specificity limit, not evidence that EOGT itself performs the downstream galactosylation. The ISS source identity is verified; unavailable original donor Methods do not overturn independent direct human evidence for this well-established process. Full Methods and Results in PMID:25488668 additionally establish catalysis by purified CHO-expressed mouse EOGT using radiolabeled UDP-GlcNAc and a fly Notch EGF20 substrate, providing a separately read primary mouse assay.

Summary: Refine the broad glycosyltransferase activity to the experimentally established protein O-acetylglucosaminyltransferase activity.

Reason: Both current InterPro mappings, IPR007657 (Glycosyltransferase 61 family) and IPR049625 (its catalytic domain), supply only the broad GO:0016757 assertion. This family also includes enzymes with other substrate specificities, so the mapping alone does not identify the acceptor. Human EOGT activity in PMID:23671640 Figure 1 and purified fly-enzyme assays in PMID:22158438 Figure 2 establish O-GlcNAc transfer to EGF-domain proteins. GO:0097363, already asserted by the IBA, combined IEA and ISS annotations, expresses that established reaction and is a verified is_a descendant of GO:0016757. Use that nonredundant specific replacement; the refinement does not imply the InterPro mapping is false.

Proposed replacements: protein O-acetylglucosaminyltransferase activity

Summary: The PAINT-inherited protein O-acetylglucosaminyltransferase activity is corroborated directly for human EOGT.

Reason: PTN001841612 carries the exact IBD term and fly/mouse donor set in the pinned PAINT data. Human, mouse and fly Eogt occupy the EOGT subfamily, and the human target descends from the asserted node without a matching loss annotation on its path. PMID:22158438 Figure 2 measures transfer from UDP-[3H]GlcNAc to folded EGF20 using affinity-purified fly Eogt; denatured/disulfide-altered acceptors and the T34A acceptor give little or no activity. PMID:23671640 Figure 1 establishes activity of the canonical human 527-residue clone in a heterologous cellular assay. The older mouse source is not needed as the sole basis for this accepted function, and no equivalence of shorter isoforms is inferred. Independently read PMID:25488668 Methods and Results corroborate this chemistry with purified mouse enzyme; the reported kinetic constants belong to that mouse preparation.

Summary: The combined orthology and reaction mapping correctly identifies protein O-GlcNAc transferase activity.

Reason: Current Rhea 48904 and 48908 describe transfer of N-acetylglucosamine from UDP-GlcNAc to protein serine or threonine, matching the alternatives in GO:0097363 and EC 2.4.1.255. The Q8BYW9 and Ensembl donor identities are verified. These reaction identifiers specify chemistry; they do not by themselves distinguish EOGT substrate/location specificity from intracellular OGT. Human activity is independently supported by PMID:23671640 Figure 1, and fly purified-enzyme specificity by PMID:22158438 Figure 2. The normal mouse 22310717 cache is abstract-only, a source-access limit rather than uncertainty about the established human catalytic function. Independently read PMID:25488668 Methods and Results corroborate donor-transfer chemistry with purified mouse enzyme; neither the mouse kinetics nor the disease-variant constructs establish equivalent activities of natural human isoforms.

Summary: The manually transferred catalytic activity agrees with direct evidence for canonical human EOGT.

Reason: The source Q8BYW9 is verified mouse Eogt, with an IDA GO:0097363 annotation citing PMID:22310717. Although that original full Methods remain inaccessible, PMID:23671640 Figure 1 and Methods independently demonstrate O-GlcNAc transfer to Notch EGF repeats by the human 527-residue clone. The tested KC347596.1 translation exactly matches Q5NDL2-1. The DYD-to-AYA comparison reduces activity rather than eliminating it; it is not a direct assay of either natural shorter product. Independently read PMID:25488668 Methods and Results provide direct mouse ortholog catalytic support. Its mouse disease-substitution constructs and truncation mimic are distinct from natural human splice products.