FAD1 (YDL045C) encodes FAD synthase (FMN adenylyltransferase, EC 2.7.7.2), the essential enzyme that adenylates FMN with ATP to form FAD, the last step of FAD biosynthesis from riboflavin. Fad1 is a 306-residue Mg2+-dependent member of the PAPS reductase (ATP pyrophosphatase) fold family, unrelated to the bacterial bifunctional RibF FAD synthetases; its crystal structure has been solved in complex with FAD. FAD1 disruption is lethal. Fad1 is mainly cytosolic, supplying FAD to extramitochondrial flavoproteins and to the mitochondrial FAD carrier Flx1, but a mitochondrial Fad1 pool has also been detected, whose abundance depends on carbon source and possibly on alternative FAD1 mRNA 3'UTRs.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003824 catalytic activity | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO from the PAPS reductase/phosphoadenosine phosphosulfate reductase domain (IPR002500) yields only the root 'catalytic activity'. True but uninformative; the specific activity is FMN adenylyltransferase. Proposed replacements: FMN adenylyltransferase activity Supporting Evidence: file:yeast/FAD1/FAD1-uniprot.txt SIMILARITY: Belongs to the PAPS reductase family. FAD1 subfamily. file:yeast/FAD1/FAD1-uniprot.txt Reaction=FMN + ATP + H(+) = FAD + diphosphate; Xref=Rhea:RHEA:17237, |
| GO:0003919 FMN adenylyltransferase activity | IBA GO_REF:0000033 | ACCEPT | Summary: FMN adenylyltransferase activity inherited from the eukaryotic FAD synthase node (human FLAD1, mouse Flad1, yeast FAD1). Correct; core MF. Supporting Evidence: PMID:7799934 S. cerevisiae transformed with FAD1 on a multicopy plasmid displays an increase in FAD synthetase activity. This is also true when the gene is expressed in Escherichia coli. file:yeast/FAD1/FAD1-uniprot.txt Reaction=FMN + ATP + H(+) = FAD + diphosphate; Xref=Rhea:RHEA:17237, |
| GO:0003919 FMN adenylyltransferase activity | IDA PMID:7799934 Cloning and characterization of FAD1, the structural gene fo... | ACCEPT | Summary: Wu et al. 1995: FAD1 on a multicopy plasmid raises FAD synthetase activity in yeast and confers activity when expressed in E. coli. Core MF. Supporting Evidence: PMID:7799934 S. cerevisiae transformed with FAD1 on a multicopy plasmid displays an increase in FAD synthetase activity. This is also true when the gene is expressed in Escherichia coli. |
| GO:0003919 FMN adenylyltransferase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined IEA (ARBA, RHEA:17237, EC 2.7.7.2); correct mapping for Fad1. Supporting Evidence: file:yeast/FAD1/FAD1-uniprot.txt Reaction=FMN + ATP + H(+) = FAD + diphosphate; Xref=Rhea:RHEA:17237, |
| GO:0003919 FMN adenylyltransferase activity | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways riboflavin/FMN/FAD pathway assigns the FMN + ATP -> FAD step (EC 2.7.7.2) to Fad1; correct. Supporting Evidence: PMID:7799934 S. cerevisiae transformed with FAD1 on a multicopy plasmid displays an increase in FAD synthetase activity. This is also true when the gene is expressed in Escherichia coli. |
| GO:0005634 nucleus | IEA GO_REF:0000044 | MARK AS OVER ANNOTATED | Summary: UniProt subcellular mapping of 'Nucleus', which UniProt itself flags as by similarity to S. pombe Fad1 (O74841, from a genome-wide GFP survey). No S. cerevisiae evidence for nuclear Fad1; nuclear FAD synthase isoforms are known in rat but not shown in yeast. Reason: The only basis is a by-similarity transfer of the nuclear signal that a genome-wide YFP screen recorded for S. pombe Fad1/nad1, a screen in which soluble metabolic enzymes often give diffuse nucleocytoplasmic signal; the S. pombe review keeps that HDA row only as non-core. S. cerevisiae FAD1 has no nuclear assay or nuclear role, and its characterised activity is extramitochondrial (PMID:7799934). A location transferred from another species' high-throughput screen is an over-annotation of this gene rather than a refuted claim. Supporting Evidence: file:yeast/FAD1/FAD1-uniprot.txt Nucleus {ECO:0000250|UniProtKB:O74841} |
| GO:0005634 nucleus | ISS GO_REF:0000024 | MARK AS OVER ANNOTATED | Summary: ISS from S. pombe Fad1 (O74841), whose nuclear assignment comes from a high-throughput GFP localisation screen. Same caveat as the IEA nucleus row: no S. cerevisiae data. Reason: Same basis as the IEA nucleus row, which this ISS row reinforces and is treated consistently with. It transfers a high-throughput S. pombe localisation (O74841) with no S. cerevisiae nuclear assay or role; the gene's characterised FAD synthetase activity is extramitochondrial (PMID:7799934). Supporting Evidence: file:yeast/FAD1/FAD1-uniprot.txt Nucleus {ECO:0000250|UniProtKB:O74841} |
| GO:0005737 cytoplasm | IDA PMID:7799934 Cloning and characterization of FAD1, the structural gene fo... | ACCEPT | Summary: Wu et al. 1995 found FAD synthetase activity extramitochondrially but not in wild-type mitochondria, supporting a cytoplasmic location. Supporting Evidence: PMID:7799934 This is supported by the absence of FAD synthetase activity in yeast mitochondria and the presence of both extramitochondrial and mitochondrial riboflavin kinase |
| GO:0005739 mitochondrion | IDA PMID:34575116 Subcellular Localization of Fad1p in Saccharomyces cerevisia... | ACCEPT | Summary: Bruni et al. 2021 detected natural Fad1 in purified mitochondria (anti-FADS antiserum) and showed membrane-potential-dependent import of tagged Fad1; abundance depends on carbon source. This contrasts with Wu et al. 1995, who found no mitochondrial FAD synthetase activity in wild type, so the mitochondrial pool is minor/conditional. Supporting Evidence: PMID:34575116 we showed that a natural Fad1p exists in yeast mitochondria and that, in its recombinant form, the protein is able, per se, to both enter mitochondria and to be destined to cytosol PMID:34575116 Thus, the Fad1p content in mitochondria seems to depend on the carbon source. |
| GO:0005739 mitochondrion | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt mapping of 'Mitochondrion', based on Bruni et al. 2021 who detected endogenous Fad1 in mitochondria (carbon-source dependent). Accept as a minor pool. Supporting Evidence: PMID:34575116 we showed that a natural Fad1p exists in yeast mitochondria and that, in its recombinant form, the protein is able, per se, to both enter mitochondria and to be destined to cytosol |
| GO:0005829 cytosol | IDA PMID:34575116 Subcellular Localization of Fad1p in Saccharomyces cerevisia... | ACCEPT | Summary: Same study shows recombinant Fad1 is also destined to the cytosol (echoforms); cytosol is the main site of FAD synthesis in yeast. Supporting Evidence: PMID:34575116 we showed that a natural Fad1p exists in yeast mitochondria and that, in its recombinant form, the protein is able, per se, to both enter mitochondria and to be destined to cytosol |
| GO:0005829 cytosol | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt mapping of 'Cytoplasm, cytosol'; the bulk of Fad1 is extramitochondrial/cytosolic. Supporting Evidence: PMID:34575116 we showed that a natural Fad1p exists in yeast mitochondria and that, in its recombinant form, the protein is able, per se, to both enter mitochondria and to be destined to cytosol PMID:7799934 This is supported by the absence of FAD synthetase activity in yeast mitochondria and the presence of both extramitochondrial and mitochondrial riboflavin kinase |
| GO:0005829 cytosol | RCA GO_REF:0000123 | ACCEPT | Summary: Pathway-default cytosol; here consistent with experimental data (Wu 1995; Bruni 2021). Supporting Evidence: PMID:7799934 This is supported by the absence of FAD synthetase activity in yeast mitochondria and the presence of both extramitochondrial and mitochondrial riboflavin kinase |
| GO:0006747 FAD biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: FAD biosynthesis inherited from the same FAD synthase node; Fad1 catalyses the final step. Core BP. Supporting Evidence: PMID:7799934 S. cerevisiae transformed with FAD1 on a multicopy plasmid displays an increase in FAD synthetase activity. This is also true when the gene is expressed in Escherichia coli. |
| GO:0006747 FAD biosynthetic process | IDA PMID:7799934 Cloning and characterization of FAD1, the structural gene fo... | ACCEPT | Summary: FAD synthetase activity of the FAD1 product directly places it in FAD biosynthesis. Supporting Evidence: PMID:7799934 S. cerevisiae transformed with FAD1 on a multicopy plasmid displays an increase in FAD synthetase activity. This is also true when the gene is expressed in Escherichia coli. |
| GO:0006747 FAD biosynthetic process | IEA GO_REF:0000041 | ACCEPT | Summary: UniPathway UPA00277 (FAD biosynthesis, FAD from FMN) mapping; correct. Supporting Evidence: file:yeast/FAD1/FAD1-uniprot.txt Reaction=FMN + ATP + H(+) = FAD + diphosphate; Xref=Rhea:RHEA:17237, |
| GO:0006747 FAD biosynthetic process | IMP PMID:7799934 Cloning and characterization of FAD1, the structural gene fo... | ACCEPT | Summary: FAD1 disruption is lethal and G178 pet mutants suppressed by FAD1 have altered FAD/FMN ratios; consistent with Fad1 being the sole FAD synthase. Supporting Evidence: PMID:7799934 FAD1 codes for an essential yeast protein, since disruption of the gene induces a lethal phenotype. |
| GO:0042727 flavin-containing compound biosynthetic process | RCA GO_REF:0000123 | TOO GENERAL | Summary: Flavin-containing compound biosynthesis inherited from the YeastPathways superpathway (riboflavin, FMN and FAD). FAD is a flavin, so correct, but less specific than FAD biosynthetic process. Supporting Evidence: PMID:7799934 S. cerevisiae transformed with FAD1 on a multicopy plasmid displays an increase in FAD synthetase activity. This is also true when the gene is expressed in Escherichia coli. |
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Download this section (compressed HTML)Q: How large and how physiologically important is the mitochondrial Fad1 pool relative to FAD import via Flx1, given that Wu et al. (1995) detected no mitochondrial FAD synthetase activity in wild type?
Q: Is there any nuclear Fad1 in S. cerevisiae (as suggested by S. pombe GFP data and rat nuclear FADS)?
Experiment: Endogenous FAD1 tagging with organelle markers on glucose vs glycerol, combined with 3'UTR-swap alleles, to test 3'UTR-dependent mitochondrial targeting.
Flavin adenine dinucleotide (FAD) synthetase; performs the second step in synthesis of FAD from riboflavin; mutation is functionally complemented by human FLAD1
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