COLGALT1 (collagen beta(1-O)galactosyltransferase 1; also known as GLT25D1) is a soluble enzyme of the endoplasmic reticulum lumen that catalyzes the first committed step of collagen O-linked glycosylation. It transfers galactose from UDP-alpha-D-galactose onto (5R)-5-hydroxy-L-lysine (hydroxylysine) residues of collagen, forming the beta(1-O)-linked Gal-O-hydroxylysine (EC 2.4.1.50). This galactose is then the acceptor for subsequent alpha1,2-glucosylation, generating the Glc(alpha1-2)Gal disaccharide that is strongly conserved across animal collagens. The enzyme belongs to glycosyltransferase family 25 (GT25), adopts a GT-A-like fold with metal-dependent (Mn2+) catalysis, and uses essential aspartate residues (D166/D168, D461/D463) for activity. COLGALT1 acts on multiple collagen types (including types I and IV) as well as the collagenous domain of mannose-binding lectin, and co-localizes in the early secretory pathway with the upstream lysyl hydroxylase PLOD3/LH3. As the predominant of two paralogous collagen galactosyltransferases (the other being COLGALT2/GLT25D2), it is broadly expressed and its loss reduces collagen glycosylation, causing intracellular collagen accumulation. Biallelic loss-of-function variants in COLGALT1 cause an autosomal recessive cerebral small vessel disease (brain small vessel disease 3).
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0050211 procollagen galactosyltransferase activity | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetically inferred procollagen galactosyltransferase activity. This is the well-established core molecular function of COLGALT1, directly demonstrated experimentally for the human enzyme. Reason: This is the core molecular function of COLGALT1, supported by direct biochemical characterization (EC 2.4.1.50) and mutagenesis. The IBA transfer is fully consistent with the experimental evidence. Supporting Evidence: PMID:19075007 Amino acid analysis of the products of GLT25D1 and GLT25D2 reactions confirmed the transfer of galactose to hydroxylysine residues. |
| GO:0005788 endoplasmic reticulum lumen | IEA GO_REF:0000044 | ACCEPT | Summary: ER lumen localization mapped from UniProt subcellular location. COLGALT1 is a soluble ER-lumen protein, directly shown by immunofluorescence and membrane-floatation assays. Reason: ER lumen is the correct site of action; this electronic mapping agrees with direct experimental localization evidence. Supporting Evidence: PMID:20470363 In agreement with the predictions our results show that GLT25D1 is directed to the ER lumen as a soluble protein and retained there. |
| GO:0050211 procollagen galactosyltransferase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Electronically inferred procollagen galactosyltransferase activity (mapped from RHEA:12637 / EC 2.4.1.50). This is the experimentally validated core function. Reason: Consistent with the experimentally demonstrated catalytic activity and EC assignment. Supporting Evidence: file:human/COLGALT1/COLGALT1-uniprot.txt EC=2.4.1.50; Reaction=(5R)-5-hydroxy-L-lysyl-[collagen] + UDP-alpha-D-galactose = (5R)-5-O-(beta-D-galactosyl)-5-hydroxy-L-lysyl-[collagen] + UDP + H(+); Xref=Rhea:RHEA:12637. |
| GO:0030199 collagen fibril organization | TAS Reactome:R-HSA-1650814 | KEEP AS NON CORE | Summary: COLGALT1 contributes to collagen biosynthesis/modification, which is upstream of collagen fibril organization in the extracellular matrix. Its direct action is intracellular hydroxylysine galactosylation, not fibril assembly itself. Reason: This is a downstream, indirect biological consequence of collagen glycosylation rather than the core molecular function. Glycosylation has been suggested to influence collagen crosslink formation and matrix organization, but the enzyme does not directly organize fibrils. Retaining as non-core captures the pathway context without overstating the role. Supporting Evidence: PMID:27402836 glycosylation may be involved in the organization of collagens in the extracellular space. Glycosylation has been suggested to regulate cross-link formation in fibrillar collagens |
| GO:0050211 procollagen galactosyltransferase activity | TAS Reactome:R-HSA-1981120 | ACCEPT | Summary: Reactome-asserted procollagen galactosyltransferase activity, the core molecular function of COLGALT1. Reason: Core molecular function, well supported by direct biochemical and mutagenesis evidence. Supporting Evidence: PMID:19075007 Amino acid analysis of the products of GLT25D1 and GLT25D2 reactions confirmed the transfer of galactose to hydroxylysine residues. |
| GO:0180062 protein O-linked glycosylation via galactose | IMP PMID:27402836 Collagen Accumulation in Osteosarcoma Cells lacking GLT25D1 ... | ACCEPT | Summary: COLGALT1 performs the galactose-transfer step of collagen O-linked glycosylation onto hydroxylysine. Its loss reduces collagen glycosylation by up to 60% in osteosarcoma cells, directly implicating it in this process. Reason: This accurately describes the biological process the enzyme directly carries out (O-linked galactosylation of hydroxylysine), supported by loss-of-function data. Supporting Evidence: PMID:27402836 Loss of GLT25D1 decreased collagen glycosylation by up to 60% |
| GO:0050211 procollagen galactosyltransferase activity | IMP PMID:27402836 Collagen Accumulation in Osteosarcoma Cells lacking GLT25D1 ... | ACCEPT | Summary: Loss of GLT25D1/COLGALT1 reduces collagen galactosylation, providing functional (mutant phenotype) support for its procollagen galactosyltransferase activity. Reason: Core molecular function with direct loss-of-function support, corroborating the biochemical characterization. Supporting Evidence: PMID:27402836 Loss of GLT25D1 decreased collagen glycosylation by up to 60% |
| GO:1904028 positive regulation of collagen fibril organization | IMP PMID:27402836 Collagen Accumulation in Osteosarcoma Cells lacking GLT25D1 ... | MARK AS OVER ANNOTATED | Summary: This annotation derives from loss-of-function studies in osteosarcoma cells, but those experiments measured collagen glycosylation, intracellular collagen accumulation, and collagen type I expression, not fibril organization. The same study reported that loss of GLT25D1 did not alter collagen folding or thermal stability. Reason: The cited paper does not demonstrate positive regulation of collagen fibril organization; it showed intracellular collagen accumulation and unchanged folding/thermal stability. Any effect on fibril organization is indirect and speculative. This regulatory term over-interprets the available evidence. Supporting Evidence: PMID:27402836 Loss of GLT25D1 decreased collagen glycosylation by up to 60% but did not alter collagen folding and thermal stability. |
| GO:0016020 membrane | HDA PMID:19946888 Defining the membrane proteome of NK cells. | MARK AS OVER ANNOTATED | Summary: This high-throughput annotation comes from a mass-spectrometry membrane-proteome screen of NK-like YTS cells. COLGALT1 is a soluble ER-lumen protein (no transmembrane domain), and the screen itself noted that a large fraction of identified proteins were not plausible integral membrane proteins but transiently associated. Reason: COLGALT1 is a soluble protein of the ER lumen, directly demonstrated by membrane-floatation and localization assays; it has a cleaved signal sequence and no transmembrane region. The membrane assignment reflects co-isolation in a proteomic screen rather than genuine membrane residence, and is contradicted by direct evidence. Supporting Evidence: PMID:20470363 The bulk of GLT25D1 is observed in the bottom fractions of the gradient, indicating the protein is not an integral membrane protein. |
| GO:0005788 endoplasmic reticulum lumen | TAS Reactome:R-HSA-1981120 | ACCEPT | Summary: ER lumen localization asserted by Reactome, consistent with direct experimental evidence that COLGALT1 is a soluble ER-lumen protein. Reason: Correct site of action, corroborated by direct localization data. Supporting Evidence: PMID:20470363 GLT25D1 is directed to the ER lumen as a soluble protein and retained there. |
| GO:0005788 endoplasmic reticulum lumen | TAS Reactome:R-HSA-8948228 | ACCEPT | Summary: ER lumen localization asserted by Reactome (COLGALT1/COLGALT2 binding lysyl-hydroxylated collagen propeptides), consistent with the soluble ER-lumen localization of the enzyme. Reason: Correct localization, corroborated by direct experimental evidence. Supporting Evidence: PMID:20470363 GLT25D1 is directed to the ER lumen as a soluble protein and retained there. |
| GO:0005788 endoplasmic reticulum lumen | TAS Reactome:R-HSA-8948231 | ACCEPT | Summary: ER lumen localization asserted by Reactome, consistent with the experimentally established soluble ER-lumen residence of COLGALT1. Reason: Correct localization, corroborated by direct experimental evidence. Supporting Evidence: PMID:20470363 GLT25D1 is directed to the ER lumen as a soluble protein and retained there. |
| GO:0005788 endoplasmic reticulum lumen | IDA PMID:20470363 The human collagen beta(1-O)galactosyltransferase, GLT25D1, ... | ACCEPT | Summary: Direct immunofluorescence and membrane-floatation evidence that COLGALT1 is a soluble protein of the ER lumen, retained via a C-terminal RDEL signal, co-localizing with PLOD3/LH3 and MBL in the early secretory pathway. Reason: This is the strongest, most direct evidence for the cellular location of COLGALT1 and represents its core site of action. Supporting Evidence: PMID:20470363 GLT25D1 is directed to the ER lumen as a soluble protein and retained there. |
| GO:0030145 manganese ion binding | IC PMID:22216269 Identification of domains and amino acids essential to the c... | NEW | Summary: Proposed annotation not present in the current GOA for COLGALT1. Reason: COLGALT1 belongs to glycosyltransferase family 25 with a GT-A-like fold and essential catalytic aspartate residues (D166/D168, D461/D463 identified by mutagenesis), characteristic of divalent-metal (Mn2+)-dependent glycosyltransferases. Mn2+ dependence is expected on family and structural grounds, although no direct experimental metal-binding annotation currently exists in GOA. Supporting Evidence: PMID:22216269 Examination of the three DXD motifs of the active GLT25D1 by site-directed mutagenesis confirmed the importance of the first (amino acids 166-168) and second motif (amino acids 461-463) for enzymatic activity, whereas the third one was dispensable PMID:22216269 The presence of DXD motifs is typical for glycosyltransferases utilizing nucleotide-activated sugars as donor substrates. Such DXD motifs are usually involved in the binding of the nucleotide diphosphate moiety through Mn 2+ chelation |
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Download this section (compressed HTML)Q: Does COLGALT1 require Mn2+ (or another divalent cation) for catalysis, and which residues coordinate the metal in the recently solved GT25 structures?
Q: To what extent are the collagen folding/secretion phenotypes of COLGALT1 loss tissue- and collagen-type-specific (e.g., type IV in cerebral vasculature vs. type I in bone), given paralogous compensation by COLGALT2?
Experiment: Use the available cryo-EM and X-ray structures (e.g., PDB 8ZGE, 9EVJ) together with metal-substitution and site-directed mutagenesis to confirm the catalytic metal requirement and map the divalent-cation coordination site.
Experiment: Generate tissue-specific or vascular COLGALT1 knockouts/knock-ins of BSVD3 variants and quantify collagen type IV glycosylation, basement membrane integrity, and small-vessel pathology to causally link enzymatic loss to disease.
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