Zebrafish glceb (glce-B) encodes D-glucuronyl C5-epimerase B, a 585-residue single-pass type II Golgi membrane enzyme. It epimerizes C5 of D-glucuronate residues that sit next to N-sulfated glucosamine in a nascent heparan sulfate or heparin chain, converting them to L-iduronate (EC 5.1.3.17) and so acting between N-sulfation and O-sulfation in the ordered chain-modification sequence (PMID:25568314). Crystal structures of the zebrafish enzyme show a stable homodimer with two catalytic sites in the C-terminal alpha-helical domains, and mutagenesis identifies Tyr468, Tyr528 and Tyr546 as essential (PMID:25568314). The iduronate it creates gives the chain conformational flexibility that underlies ligand binding and cell signalling. Zebrafish carries two paralogs of the single human GLCE gene, glce-A and glce-B, both maternally deposited and functionally interchangeable in overexpression assays; raising glce levels ventralizes embryos and potentiates Bmp2b, while knockdown dorsalizes them, placing the epimerase upstream of BMP-dependent dorsoventral patterning (PMID:16156897).
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0015012 heparan sulfate proteoglycan biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from the GLCE node for heparan sulfate proteoglycan biosynthesis, with fly, worm, mouse and human donors. C5-epimerization is one of the ordered chain-modification steps (after N-deacetylation/N-sulfation, before O-sulfation) that build a mature HS chain (PMID:25568314). Reason: Participation is direct: glceb performs one of the chemical steps of the pathway rather than being required for it from outside. The node placement matches a family that does nothing else. Core process. Supporting Evidence: PMID:25568314 A heparan sulfate chain is synthesized in vivo by several steps: tetrasaccharide linkage formation, chain elongation, N -deacetylation/ N -sulfation, epimerization, and O -sulfation ( Fig. file:DANRE/glceb/glceb-uniprot.txt PATHWAY: Glycan metabolism; heparan sulfate biosynthesis. |
| GO:0047464 heparosan-N-sulfate-glucuronate 5-epimerase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from the GLCE node (PTN002663262) seeded by mouse Glce, human GLCE (O94923), C. elegans hse-5 (O18756) and this zebrafish gene. The family has a single, conserved activity and the target carries its own IDA for the same term. Reason: The node is a one-function family and the target is inside it; appearance of the zebrafish gene in its own WITH/FROM is the expected mark that experimental grounding exists on the target (PMID:25568314), not circularity. Same action as the IDA row. Supporting Evidence: PMID:25568314 d -Glucuronyl C5-epimerase (Glce) is a key enzyme in HS/heparin synthesis, converting d -glucuronic acid (GlcA) to l -iduronic acid (IdoA) by C5 epimerization at the polymer level ( 4 ) ( Fig. file:DANRE/glceb/glceb-uniprot.txt Converts D-glucuronic acid residues adjacent to N-sulfate |
| GO:0005794 Golgi apparatus | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: IBA from a broader node (PTN000995293) placing the protein in the Golgi apparatus, the parent of the Golgi membrane term carried by the IEA and ISS rows. Reason: Correct but redundant: the specific child GO:0000139 Golgi membrane is annotated from UniProt and is the informative statement, since glceb is a single-pass type II membrane protein rather than a lumenal Golgi resident. Kept as non-core rather than modified, because the parent term makes no claim the child does not already cover. Supporting Evidence: file:DANRE/glceb/glceb-uniprot.txt SUBCELLULAR LOCATION: Golgi apparatus membrane |
| GO:0000139 Golgi membrane | IEA GO_REF:0000044 | ACCEPT | Summary: IEA from the UniProt subcellular-location vocabulary (SL-0134): Golgi apparatus membrane, single-pass type II membrane protein. The Golgi is where HS chains are polymerized and modified, so it is where the epimerase must act. Reason: Placing the enzyme in the Golgi membrane follows from the pathway it acts in: the substrate is a nascent HS chain on a proteoglycan core protein traversing the Golgi (PMID:25568314). Accepted as the location of the core activity; the assignment is by similarity to rat Glce, with no zebrafish imaging. Supporting Evidence: file:DANRE/glceb/glceb-uniprot.txt SUBCELLULAR LOCATION: Golgi apparatus membrane PMID:25568314 A heparan sulfate chain is synthesized in vivo by several steps: tetrasaccharide linkage formation, chain elongation, N -deacetylation/ N -sulfation, epimerization, and O -sulfation ( Fig. |
| GO:0015012 heparan sulfate proteoglycan biosynthetic process | IEA GO_REF:0000120 | ACCEPT | Summary: Automated annotation from InterPro IPR039721 and UniPathway UPA00756 for the same HS biosynthetic process. Reason: Same claim as the IBA row and supported the same way: the epimerization step is part of HS chain maturation (PMID:25568314), and UniProt records the pathway for this entry. Supporting Evidence: file:DANRE/glceb/glceb-uniprot.txt PATHWAY: Glycan metabolism; heparan sulfate biosynthesis. PMID:25568314 A heparan sulfate chain is synthesized in vivo by several steps: tetrasaccharide linkage formation, chain elongation, N -deacetylation/ N -sulfation, epimerization, and O -sulfation ( Fig. |
| GO:0047464 heparosan-N-sulfate-glucuronate 5-epimerase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined automated annotation from InterPro IPR039721 (GLCE family), RHEA:20197 and EC 5.1.3.17 for the same epimerase activity. Reason: The InterPro family is GLCE-specific rather than a broad fold signature, and the Rhea reaction is the one curated on this UniProt entry from the zebrafish structure paper (PMID:25568314). Consistent with the IDA and IBA rows. Supporting Evidence: file:DANRE/glceb/glceb-uniprot.txt Converts D-glucuronic acid residues adjacent to N-sulfate PMID:25568314 d -Glucuronyl C5-epimerase (Glce) is a key enzyme in HS/heparin synthesis, converting d -glucuronic acid (GlcA) to l -iduronic acid (IdoA) by C5 epimerization at the polymer level ( 4 ) ( Fig. |
| GO:0030210 heparin proteoglycan biosynthetic process | IEA GO_REF:0000041 | KEEP AS NON CORE | Summary: UniPathway IEA (UPA00862) for heparin proteoglycan biosynthesis. Heparin is the highly sulfated, IdoA-rich relative of HS made in connective-tissue mast cells; UniProt lists the enzyme in both pathways because the same epimerization step occurs in each, and the zebrafish structure was solved with a heparin hexasaccharide product (PMID:25568314). Reason: The chemistry is identical and the row is not wrong, but heparin production is a specialized cell-type-restricted output and there is no zebrafish evidence that glceb serves it; the demonstrated context is HS chain maturation. Kept as non-core rather than accepted alongside GO:0015012. Supporting Evidence: file:DANRE/glceb/glceb-uniprot.txt PATHWAY: Glycan metabolism; heparin biosynthesis. file:DANRE/glceb/glceb-uniprot.txt Converts D-glucuronic acid residues adjacent to N-sulfate |
| GO:0042802 identical protein binding | IPI PMID:25568314 Structural and functional study of D-glucuronyl C5-epimerase... | KEEP AS NON CORE | Summary: IPI from the zebrafish Glce crystal structures (PMID:25568314), which were solved in a stable dimer conformation, with each dimer containing two catalytic sites in the C-terminal alpha-helical domains. Bacterial Glce is a dimer too, supporting that the dimer is the functional unit. Reason: Self-association is real and structurally meaningful here - the catalytic clefts are formed in the dimer and a long HS chain may thread through both sites (PMID:25568314) - so this is not an uninformative binding annotation. But the informative molecular function is already recorded as GO:0047464; homodimerization is the quaternary arrangement that supports it, not a separate activity. Kept as non-core. Supporting Evidence: PMID:25568314 Here, we report that zebrafish Glce has a dimeric structure. PMID:25568314 A Glce dimer contains two catalytic sites, each at a positively charged cleft in C-terminal Ξ±-helical domains binding one negatively charged hexasaccharide. PMID:25568314 Together, these findings strongly support the concept that Glce functions as a dimer. file:DANRE/glceb/glceb-uniprot.txt SUBUNIT: Homodimer. |
| GO:0000139 Golgi membrane | ISS GO_REF:0000024 | ACCEPT | Summary: Manual ISS transfer of the Golgi membrane location from rat Glce (UniProtKB:Q9EPS3), the same donor that underlies the UniProt SUBCELLULAR LOCATION line. Reason: Same claim and same basis as the UniProt-vocabulary IEA row, and the type II membrane topology is conserved in the zebrafish sequence. Accepted for consistency; a zebrafish localization experiment is listed under suggested_experiments. Supporting Evidence: file:DANRE/glceb/glceb-uniprot.txt SUBCELLULAR LOCATION: Golgi apparatus membrane |
| GO:0047464 heparosan-N-sulfate-glucuronate 5-epimerase activity | IDA PMID:25568314 Structural and functional study of D-glucuronyl C5-epimerase... | ACCEPT | Summary: IDA from the crystallographic and mutagenesis study of zebrafish Glce (PMID:25568314): apo and heparin-hexasaccharide-bound structures were solved and Tyr468, Tyr528 and Tyr546 shown by mutagenesis to be required for activity. The enzyme epimerizes C5 of D-glucuronate adjacent to N-sulfated glucosamine, giving L-iduronate in the growing chain. Deep research adds that the enzyme only acts on chain regions already N-sulfated at the glucosamine units, which is what couples this step to the preceding N-deacetylation/N-sulfation. Reason: Direct structural and enzymatic evidence on the zebrafish protein itself, with the catalytic residues identified by mutation (PMID:25568314); this is the reaction UniProt records for the entry (RHEA:20197, EC 5.1.3.17). The core molecular function. Supporting Evidence: PMID:25568314 d -Glucuronyl C5-epimerase (Glce) is a key enzyme in HS/heparin synthesis, converting d -glucuronic acid (GlcA) to l -iduronic acid (IdoA) by C5 epimerization at the polymer level ( 4 ) ( Fig. PMID:25568314 Based on the structural and mutagenesis studies, three tyrosine residues, Tyr 468 , Tyr 528 , and Tyr 546 , in the active site were found to be crucial for the enzymatic activity. file:DANRE/glceb/glceb-uniprot.txt Converts D-glucuronic acid residues adjacent to N-sulfate file:DANRE/glceb/glceb-deep-research-falcon.md GLCE preferentially recognizes HS regions where adjacent glucosamine residues are **N-sulfated** (GlcNS) |
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Download this section (compressed HTML)Q: Do zebrafish glceb (glce-B) and glcea (glce-A) have distinct or redundant roles in HS fine structure and morphogen signaling, given their overlapping expression and similar overexpression/knockdown phenotypes?
Q: Does the reported physical association of Glce with 2-O- and 6-O-sulfotransferases (the proposed 'GAGosome') operate in zebrafish, and is it required for proper IdoA/O-sulfation patterning in vivo?
Q: Is there any division of labour between glceb and glcea in vivo, given that their overexpression phenotypes are indistinguishable (PMID:16156897) and no single-gene mutant has been reported?
Experiment: Generate glceb loss-of-function mutants (CRISPR) and assay HS disaccharide composition (IdoA content) together with dorsoventral patterning markers to define the in vivo requirement for glceb specifically versus glcea.
Experiment: Image tagged or endogenous Glceb against Golgi markers in zebrafish embryos, which would convert the current ISS/IEA location assignment into direct evidence.
Hypothesis: Zebrafish glceb protein resides in the Golgi membrane, as inferred from rat Glce.
Type: imaging
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