Sec59 (SPCC63.10c) is the fission yeast dolichol kinase, a 504-residue polytopic endoplasmic-reticulum membrane protein of the polyprenol kinase family. It catalyzes CTP-dependent phosphorylation of free dolichol to dolichyl monophosphate (Dol-P), the terminal step of de novo Dol-P synthesis. Dol-P is the lipid carrier on which the N-glycan precursor is assembled and the acceptor for formation of dolichyl-phosphate-mannose and dolichyl-phosphate-glucose, so Sec59 supplies the carrier used by N-linked glycosylation, protein O-mannosylation and GPI-anchor biosynthesis. Its function is inferred from membership in the same PANTHER dolichol kinase subfamily as biochemically and genetically characterized budding-yeast Sec59 and human DOLK; the pombe protein itself has been localized to the ER in a proteome-wide tagging screen but has not been assayed enzymatically.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004168 dolichol kinase activity | IBA GO_REF:0000033 | ACCEPT | Summary: Dolichol kinase activity inherited from the eukaryotic dolichol kinase node PTN000324722. Reason: The PAINT curator placed dolichol kinase activity at PTN000324722 (eukaryote-level node), seeded by experimental annotations of S. cerevisiae SEC59 (SGD:S000004615), human DOLK (Q9UPQ8) and a trypanosome dolichol kinase (Q38DD5). Q9Y7T6 sits in the same PANTHER subfamily (PTHR13205:SF15, DOLICHOL KINASE) as SEC59 and DOLK, has the full-length polytopic architecture of these enzymes, and there is no evidence of loss of function in the fission yeast lineage. In S. cerevisiae the sec59 mutant lacks dolichol kinase activity and overexpression raises it, establishing the donor function. This is the core molecular function of the protein. Supporting Evidence: file:interpro/panther/PTHR13205/PTHR13205-entries.csv sec59,504,PTHR13205:SF15,DOLICHOL KINASE file:interpro/panther/PTHR13205/PTHR13205-entries.csv SEC59,519,PTHR13205:SF15,DOLICHOL KINASE PMID:1323123 When sec59 cells were transformed with a plasmid that overexpresses the wild-type gene, dolichol kinase activity increased 10-fold over wild-type levels. PMID:12213788 Dolichol kinase (DK) catalyzes the CTP-mediated phosphorylation of dolichol in eukaryotic cells, the terminal step in dolichyl monophosphate (Dol-P) biosynthesis de novo. |
| GO:0004168 dolichol kinase activity | IEA GO_REF:0000120 | ACCEPT | Summary: EC 2.7.1.108 / RHEA:13133 mapping to dolichol kinase activity. Reason: The EC/Rhea assignment on the UniProt record is itself an ortholog-based (ECO:0000250 from P20048) curation and agrees with the PANTHER SF15 placement and the IBA for the same term. The reaction (dolichol + CTP = dolichyl phosphate + CDP) is the established activity of the subfamily. Supporting Evidence: file:SCHPO/sec59/sec59-uniprot.txt CC -!- FUNCTION: Catalyzes CTP-mediated phosphorylation of dolichol, the PMID:12213788 Dolichol kinase (DK) catalyzes the CTP-mediated phosphorylation of dolichol in eukaryotic cells, the terminal step in dolichyl monophosphate (Dol-P) biosynthesis de novo. |
| GO:0004168 dolichol kinase activity | ISS GO_REF:0000024 | ACCEPT | Summary: Curator transfer from experimentally characterized S. cerevisiae Sec59 (P20048). Reason: P20048 is in the same PANTHER subfamily (PTHR13205:SF15) and its dolichol kinase function is experimentally established (sec59 mutant membranes deficient in dolichol kinase and Dol-P; complementation by human DOLK). The transfer is phylogenetically sound and consistent with the IBA. Supporting Evidence: file:SCHPO/sec59/sec59-P20048-ortholog-uniprot.txt CC -!- FUNCTION: Catalyzes CTP-mediated phosphorylation of dolichol, the PMID:1323123 However, dolichol kinase activity was greatly decreased. |
| GO:0005783 endoplasmic reticulum | HDA PMID:16823372 ORFeome cloning and global analysis of protein localization ... | MODIFY | Summary: Proteome-wide YFP tagging placed Sec59 in the ER. Reason: The HDA observation is the only direct pombe evidence and is sound, but the protein has ten predicted transmembrane segments and UniProt curates the same study as ER membrane localization. The specific part_of descendant GO:0005789 endoplasmic reticulum membrane, which the gene already carries by IBA and IEA, is the appropriate term. Proposed replacements: endoplasmic reticulum membrane Supporting Evidence: file:SCHPO/sec59/sec59-uniprot.txt CC -!- SUBCELLULAR LOCATION: Endoplasmic reticulum membrane PMID:16823372 we determined the localization of 4,431 proteins, corresponding to approximately 90% of the fission yeast proteome, by tagging each ORF with the yellow fluorescent protein. |
| GO:0005789 endoplasmic reticulum membrane | IBA GO_REF:0000033 | ACCEPT | Summary: ER membrane location inherited at PTN000324722, consistent with the pombe ER localization. Reason: Dolichol kinases are polytopic ER membrane proteins (S. cerevisiae SEC59, human DOLK, an Arabidopsis member are the donors). The target has ten predicted transmembrane helices and was observed at the ER in pombe, so the inherited location is independently corroborated. Supporting Evidence: file:SCHPO/sec59/sec59-uniprot.txt CC -!- SUBCELLULAR LOCATION: Endoplasmic reticulum membrane file:SCHPO/sec59/sec59-uniprot.txt FT TRANSMEM 56..76 |
| GO:0005789 endoplasmic reticulum membrane | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular-location mapping to ER membrane. Reason: The UniProt subcellular location is backed by the pombe localization study (ECO:0000269, PubMed:16823372) together with the multipass topology; same claim as the IBA row. Supporting Evidence: file:SCHPO/sec59/sec59-uniprot.txt CC -!- SUBCELLULAR LOCATION: Endoplasmic reticulum membrane |
| GO:0043048 dolichyl monophosphate biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: Sec59 catalyzes the terminal step of de novo Dol-P synthesis. Reason: Dolichol phosphorylation is the final step of de novo dolichyl monophosphate formation, so the enzyme does the work of this process directly. In S. cerevisiae sec59 cells Dol-P falls to under 10% of wild type at the restrictive temperature while other Dol-P pathway enzymes are normal. The PAINT node is the same one carrying the kinase activity. Supporting Evidence: PMID:1323123 Quantitative measurements of dolichyl phosphate in sec59 cells showed that the levels were decreased to 48% of wild type at the permissive temperature and to less than 10% at the restrictive temperature. PMID:12213788 Dolichol kinase (DK) catalyzes the CTP-mediated phosphorylation of dolichol in eukaryotic cells, the terminal step in dolichyl monophosphate (Dol-P) biosynthesis de novo. |
| GO:0180047 dolichol phosphate mannose biosynthetic process | IC GO_REF:0000111 | KEEP AS NON CORE | Summary: Sec59 catalyzes an upstream step (Dol-P formation) of the pathway leading to Dol-P-mannose. Reason: GO:0180047 is defined as the chemical reactions and pathways resulting in formation of dolichol phosphate mannose, and the dolichyl phosphate made by Sec59 becomes the lipid moiety of Dol-P-Man, so Sec59 catalyzes a step of this pathway rather than merely being required for it. In S. cerevisiae sec59 membranes, Dol-P-Man synthesis is depressed unless exogenous Dol-P is supplied. The PomBase IC is therefore sound, but Dol-P-Man is only one of several Dol-P-dependent products (also Dol-PP-oligosaccharide and Dol-P-glucose), so the term is kept as non-core; the direct core process is dolichyl monophosphate biosynthesis. Supporting Evidence: PMID:1323123 made normal amounts of dolichyl phosphate mannose when exogenous dolichyl phosphate was supplied, but dolichyl phosphate mannose synthesis was severely depressed in the absence of exogenous dolichyl phosphate. |
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Download this section (compressed HTML)Q: Does recombinant or microsomal S. pombe Sec59 show CTP-dependent dolichol kinase activity, and what is its preference among the dolichol chain lengths present in fission yeast?
Experiment: Complement an S. cerevisiae sec59-1 temperature-sensitive mutant with S. pombe sec59 and measure dolichol kinase activity ([gamma-32P]CTP and dolichol) in membranes, plus Dol-P levels and CPY glycosylation.
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