RNR2 (CRT6, YJL026W) encodes the radical-bearing small (R2, beta) subunit of the class Ia ribonucleotide reductase of Saccharomyces cerevisiae. Rnr2 binds the di-iron centre that generates and stores the tyrosyl radical required to initiate nucleotide reduction in the large subunit (Rnr1, or Rnr3 after DNA damage). In yeast the functional small subunit is an obligate 1:1 heterodimer of Rnr2 with the catalytically inactive paralog Rnr4, which lacks iron ligands but is needed for Rnr2 folding, cofactor assembly and docking with the large subunit; Rnr2 homodimers are essentially inactive. The holoenzyme reduces ADP, GDP, CDP and UDP to the deoxyribonucleoside diphosphates, the rate-limiting step of dNTP synthesis. RNR2 is essential. The Rnr2-Rnr4 heterodimer is held in the nucleus outside S phase by Wtm1 and imported by Dif1, and is released to the cytoplasm to join Rnr1 in S phase or after DNA damage under Mec1-Rad53-Dun1 checkpoint control; RNR2 transcription is also damage-inducible.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004748 ribonucleoside-diphosphate reductase activity, thioredoxin disulfide as acceptor | IDA PMID:10535923 Purification of ribonucleotide reductase subunits Y1, Y2, Y3... | ACCEPT | Summary: Nguyen et al. purified yeast-expressed (His)6-Rnr2 (Y2) with measurable specific activity, which rose markedly when Y4, Fe2+ and O2 were added, establishing Rnr2 as the radical-bearing catalytic small subunit. Reason: Direct activity measurement on the yeast protein; Rnr2 carries the di-iron tyrosyl radical cofactor. Supporting Evidence: PMID:10535923 RNR2 contains the diiron-tyrosyl radical (Y.) cofactor essential for the reduction process. |
| GO:0004748 ribonucleoside-diphosphate reductase activity, thioredoxin disulfide as acceptor | IDA PMID:10716984 Yeast ribonucleotide reductase has a heterodimeric iron-radi... | ACCEPT | Summary: Chabes et al. reconstituted Rnr1 + Rnr2/Rnr4 and measured 2,250 nmol dCDP/min/mg for the heterodimer-bound Rnr2; the contributes_to qualifier reflects that activity needs the large subunit. Reason: Direct biochemical evidence; core. Supporting Evidence: PMID:10716984 The specific activity of the Rnr2p complexed with Rnr4p is 2,250 nmol deoxycytidine 5'-diphosphate formed per min per mg, whereas the homodimer of Rnr2p shows no activity. PMID:10716984 the Rnr2p/Rnr4p heterodimer is the active form of the yeast RNR small subunit |
| GO:0004748 ribonucleoside-diphosphate reductase activity, thioredoxin disulfide as acceptor | IEA GO_REF:0000120 | ACCEPT | Summary: RHEA:23252 / EC 1.17.4.1 mapping from UniProt catalytic activity for the RNR small subunit family. Correct for the radical-bearing subunit. Reason: Consistent with experimental evidence. Supporting Evidence: file:yeast/RNR2/RNR2-uniprot.txt corresponding ribonucleotides. RNR2 provides the diiron-tyrosyl radical |
| GO:0004748 ribonucleoside-diphosphate reductase activity, thioredoxin disulfide as acceptor | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA (PWY-7222-1) attaches the RNR1-4 complexes to GDP reduction. Correct for Rnr2. Reason: Consistent with experimental evidence. Supporting Evidence: PMID:10716984 the Rnr2p/Rnr4p heterodimer is the active form of the yeast RNR small subunit |
| GO:0005515 protein binding | IPI PMID:10716984 Yeast ribonucleotide reductase has a heterodimeric iron-radi... | REMOVE | Summary: IPI Rnr2-Rnr1 (Chabes et al. 2000; Rnr2/Rnr4 cosediments with Rnr1 as an alpha2-beta-beta' complex). Reason: Bare protein binding to Rnr1 records the alpha-beta holoenzyme contact. It does not dispute the interaction, but the information is better captured by RNR complex membership (GO:0005971) and contributes_to RNR activity. Supporting Evidence: PMID:10716984 In the presence of Rnr1p, both polypeptides of the Rnr2p/Rnr4p heterodimer cosediment at 9.7 S as expected for an alpha(2)betabeta' heterotetramer |
| GO:0005515 protein binding | IPI PMID:10716984 Yeast ribonucleotide reductase has a heterodimeric iron-radi... | MODIFY | Summary: IPI Rnr2-Rnr4 (Chabes et al. 2000): stable 1:1 beta-beta' heterodimer that is the only active form of the small subunit. Reason: Informative replacement is protein heterodimerization activity (already annotated by IDA). Proposed replacements: protein heterodimerization activity Supporting Evidence: PMID:10716984 This complex sediments at 5.6 S as a betabeta' heterodimer in a sucrose gradient. |
| GO:0005515 protein binding | IPI PMID:11526233 Structure of the yeast ribonucleotide reductase Y2Y4 heterod... | MODIFY | Summary: IPI Rnr2-Rnr4 from the Y2Y4 heterodimer crystal structure. Reason: Replace bare binding with protein heterodimerization activity. Proposed replacements: protein heterodimerization activity Supporting Evidence: PMID:11526233 The heterodimer interface is similar to the mouse R2 homodimer interface in size and interacting residues |
| GO:0005515 protein binding | IPI PMID:16429126 Proteome survey reveals modularity of the yeast cell machine... | REMOVE | Summary: Rnr2-Rnr4 co-purification in the Gavin et al. 2006 proteome-wide TAP survey (PMID:16429126). Reason: High-throughput interaction data; bare 'protein binding' carries no functional information and the Rnr2-Rnr4/Rnr1 associations are captured by complex membership and heterodimerization annotations. Supporting Evidence: PMID:10716984 This complex sediments at 5.6 S as a betabeta' heterodimer in a sucrose gradient. |
| GO:0005515 protein binding | IPI PMID:16432237 Nuclear localization of the Saccharomyces cerevisiae ribonuc... | MODIFY | Summary: IPI Rnr2-Rnr4 from Zhang et al. 2006 (Rnr2/Rnr4 co-purify with Wtm1/Wtm2). Reason: The interaction is the beta-beta' heterodimer; replace with protein heterodimerization activity. Proposed replacements: protein heterodimerization activity Supporting Evidence: PMID:16432237 Wtm1 and its paralog Wtm2 are both nuclear proteins that are in the same protein complex with betabeta'. |
| GO:0005515 protein binding | IPI PMID:16537480 Structures of eukaryotic ribonucleotide reductase I define g... | REMOVE | Summary: IPI Rnr2-Rnr1 from Xu et al. 2006: an Rnr2 C-terminal peptide binds Rnr1 in the crystal structure, defining the alpha-beta docking site. Reason: Bare protein binding to Rnr1 records the alpha-beta holoenzyme contact. It does not dispute the interaction, but the information is better captured by RNR complex membership (GO:0005971) and contributes_to RNR activity. Supporting Evidence: PMID:16537480 Rnr2, the beta subunit (in yeast, a heterodimer of Rnr2 and Rnr4), houses the diferric-tyrosyl radical crucial for catalysis. |
| GO:0005515 protein binding | IPI PMID:21336276 Structural basis for allosteric regulation of human ribonucl... | REMOVE | Summary: IPI Rnr2-Rnr1 from Fairman et al. 2011 (EM of the dATP-induced alpha6-beta-beta' holocomplex). Reason: Bare protein binding to Rnr1 records the alpha-beta holoenzyme contact. It does not dispute the interaction, but the information is better captured by RNR complex membership (GO:0005971) and contributes_to RNR activity. Supporting Evidence: PMID:21336276 Site-directed mutagenesis and functional assays confirm that hexamerization is a prerequisite for inhibition by dATP. |
| GO:0005515 protein binding | IPI PMID:21336276 Structural basis for allosteric regulation of human ribonucl... | MODIFY | Summary: IPI Rnr2-Rnr4 from Fairman et al. 2011, which used the yeast beta-beta' heterodimer in holocomplex reconstructions. Reason: Replace with protein heterodimerization activity. Proposed replacements: protein heterodimerization activity Supporting Evidence: PMID:10716984 This complex sediments at 5.6 S as a betabeta' heterodimer in a sucrose gradient. |
| GO:0005515 protein binding | IPI PMID:37968396 The social and structural architecture of the yeast protein ... | REMOVE | Summary: Rnr2-Rnr1 interaction from the proteome-scale interactome of Michaelis et al. 2023. Reason: High-throughput interaction data; bare 'protein binding' carries no functional information and the Rnr2-Rnr4/Rnr1 associations are captured by complex membership and heterodimerization annotations. Supporting Evidence: PMID:10716984 In the presence of Rnr1p, both polypeptides of the Rnr2p/Rnr4p heterodimer cosediment at 9.7 S as expected for an alpha(2)betabeta' heterotetramer |
| GO:0005515 protein binding | IPI PMID:37968396 The social and structural architecture of the yeast protein ... | REMOVE | Summary: Rnr2-Rnr4 interaction from the proteome-scale interactome of Michaelis et al. 2023. Reason: High-throughput interaction data; bare 'protein binding' carries no functional information and the Rnr2-Rnr4/Rnr1 associations are captured by complex membership and heterodimerization annotations. Supporting Evidence: PMID:10716984 This complex sediments at 5.6 S as a betabeta' heterodimer in a sucrose gradient. |
| GO:0005634 nucleus | IDA PMID:12732713 Subcellular localization of yeast ribonucleotide reductase r... | ACCEPT | Summary: Yao et al. immunofluorescence: Rnr2 and Rnr4 are predominantly nuclear in unperturbed G1/G2 cells. Nuclear sequestration (Wtm1 anchoring, Dif1 import) is a regulatory storage site rather than where reduction occurs. Reason: Well-supported location; regulatory storage compartment. Supporting Evidence: PMID:12732713 Rnr2 and Rnr4 are predominantly present in the nucleus |
| GO:0005634 nucleus | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt SubCell mapping (Nucleus), based on PMID:12732713 and PMID:18851834. Correct. Reason: Correct. Supporting Evidence: file:yeast/RNR2/RNR2-uniprot.txt SUBCELLULAR LOCATION: Nucleus {ECO:0000269|PubMed:12732713, |
| GO:0005737 cytoplasm | IDA PMID:12732713 Subcellular localization of yeast ribonucleotide reductase r... | ACCEPT | Summary: Yao et al.: Rnr2/Rnr4 redistribute to the cytoplasm in S phase and after genotoxic stress, where they join Rnr1 to form the active enzyme. Reason: Cytoplasm is the site of the active holoenzyme. Supporting Evidence: PMID:12732713 Under genotoxic stress, Rnr2 and Rnr4 become redistributed to the cytoplasm in a checkpoint-dependent manner. |
| GO:0005737 cytoplasm | IDA PMID:9315670 Identification of RNR4, encoding a second essential small su... | ACCEPT | Summary: Huang and Elledge 1997 (abstract only in cache) reported RNR2/RNR4 co-immunoprecipitation and localisation data; cytoplasmic presence is consistent with later IDA evidence. Reason: Deferring to curator who read the full text; consistent with PMID:12732713. Supporting Evidence: PMID:9315670 RNR2 and RNR4 may be present in the same reductase complex in vivo, since they coimmunoprecipitate from cell extracts. PMID:12732713 Under genotoxic stress, Rnr2 and Rnr4 become redistributed to the cytoplasm in a checkpoint-dependent manner. |
| GO:0005737 cytoplasm | NAS PMID:16537480 Structures of eukaryotic ribonucleotide reductase I define g... | ACCEPT | Summary: NAS from the Rnr1 structure paper; consistent with the cytoplasmic active complex. Reason: Consistent with IDA. Supporting Evidence: PMID:12732713 Under genotoxic stress, Rnr2 and Rnr4 become redistributed to the cytoplasm in a checkpoint-dependent manner. |
| GO:0005737 cytoplasm | NAS PMID:18851834 Dif1 is a DNA-damage-regulated facilitator of nuclear import... | ACCEPT | Summary: NAS from Lee et al. 2008 (Dif1): Rnr2-Rnr4 enters the cytoplasm to bind Rnr1 in S phase or after damage. Reason: Consistent with IDA. Supporting Evidence: PMID:18851834 In response to S phase or DNA damage, Rnr2-Rnr4 enters the cytoplasm to bind Rnr1, forming an active complex. |
| GO:0005737 cytoplasm | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways RCA places RNR activity in cytoplasm (is_active_in). Correct for the active Rnr1-Rnr2-Rnr4 holoenzyme. Reason: Correct. Supporting Evidence: PMID:18851834 In response to S phase or DNA damage, Rnr2-Rnr4 enters the cytoplasm to bind Rnr1, forming an active complex. |
| GO:0005971 ribonucleoside-diphosphate reductase complex | IDA PMID:10716984 Yeast ribonucleotide reductase has a heterodimeric iron-radi... | ACCEPT | Summary: Chabes et al. 2000: Rnr2/Rnr4 heterodimer cosediments with Rnr1 as the alpha2-beta-beta' holoenzyme. Reason: Core complex membership. Supporting Evidence: PMID:10716984 In the presence of Rnr1p, both polypeptides of the Rnr2p/Rnr4p heterodimer cosediment at 9.7 S as expected for an alpha(2)betabeta' heterotetramer |
| GO:0005971 ribonucleoside-diphosphate reductase complex | IDA PMID:11526233 Structure of the yeast ribonucleotide reductase Y2Y4 heterod... | ACCEPT | Summary: Voegtli et al. 2001 crystal structure of the Y2Y4 heterodimer (the small-subunit part of the complex). Reason: Core complex membership. Supporting Evidence: PMID:11526233 The heterodimer interface is similar to the mouse R2 homodimer interface in size and interacting residues |
| GO:0005971 ribonucleoside-diphosphate reductase complex | IPI PMID:10716984 Yeast ribonucleotide reductase has a heterodimeric iron-radi... | ACCEPT | Summary: IPI from Chabes et al. 2000 (Rnr2 with Rnr4 and Rnr1). Reason: Core complex membership. Supporting Evidence: PMID:10716984 In the presence of Rnr1p, both polypeptides of the Rnr2p/Rnr4p heterodimer cosediment at 9.7 S as expected for an alpha(2)betabeta' heterotetramer |
| GO:0008198 ferrous iron binding | IDA PMID:11526233 Structure of the yeast ribonucleotide reductase Y2Y4 heterod... | ACCEPT | Summary: Soaking Y2Y4 crystals with Fe(II) produced density at both Fe1 and Fe2 sites of Rnr2 (not Rnr4). Ferrous iron assembles the diferric-tyrosyl radical cofactor, so this cofactor binding is intrinsic to Rnr2's catalytic role. Reason: Direct structural evidence; Rnr2 is the iron-binding subunit. Supporting Evidence: PMID:11526233 Treatment of the crystals with Fe(II) results in difference electron density consistent with formation of a diiron center. |
| GO:0008270 zinc ion binding | IDA PMID:11526233 Structure of the yeast ribonucleotide reductase Y2Y4 heterod... | MARK AS OVER ANNOTATED | Summary: A single Zn(II) occupies the Fe2 position in the as-crystallised Rnr2 active site. This is a metal substitution in the iron site of the purified protein; there is no evidence that zinc is a physiological ligand of Rnr2, whose function requires a di-iron centre. Reason: Non-physiological metal occupancy observed in the crystal; overstates Rnr2 function. Supporting Evidence: PMID:11526233 A single metal ion, assigned as Zn(II), occupies the Fe2 position in the Y2 active site. PMID:11526233 Treatment of the crystals with Fe(II) results in difference electron density consistent with formation of a diiron center. |
| GO:0009263 deoxyribonucleotide biosynthetic process | IDA PMID:10535923 Purification of ribonucleotide reductase subunits Y1, Y2, Y3... | ACCEPT | Summary: Nguyen et al. activity of purified Rnr2 in NDP reduction. Reason: Core process. Supporting Evidence: PMID:10535923 RNR2 contains the diiron-tyrosyl radical (Y.) cofactor essential for the reduction process. |
| GO:0009263 deoxyribonucleotide biosynthetic process | IDA PMID:10716984 Yeast ribonucleotide reductase has a heterodimeric iron-radi... | ACCEPT | Summary: Chabes et al. reconstituted RNR activity with Rnr2/Rnr4. Reason: Core process. Supporting Evidence: PMID:10716984 The specific activity of the Rnr2p complexed with Rnr4p is 2,250 nmol deoxycytidine 5'-diphosphate formed per min per mg, whereas the homodimer of Rnr2p shows no activity. |
| GO:0009263 deoxyribonucleotide biosynthetic process | IDA PMID:5459124 Ribonucleotide reductase from Saccharomyces cerevisiae. | ACCEPT | Summary: Vitols et al. 1970 yeast RNR activity (no abstract cached); process correct for Rnr2. Reason: Deferring to curator; process independently supported by PMID:10716984. Supporting Evidence: PMID:10716984 The specific activity of the Rnr2p complexed with Rnr4p is 2,250 nmol deoxycytidine 5'-diphosphate formed per min per mg, whereas the homodimer of Rnr2p shows no activity. |
| GO:0009263 deoxyribonucleotide biosynthetic process | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro IPR000358/IPR033909 (RNR small subunit) mapping. Correct. Reason: Correct. Supporting Evidence: file:yeast/RNR2/RNR2-uniprot.txt corresponding ribonucleotides. RNR2 provides the diiron-tyrosyl radical |
| GO:0016491 oxidoreductase activity | IEA GO_REF:0000002 | MODIFY | Summary: InterPro IPR012348 (ferritin-like RNR small subunit fold) mapping to generic oxidoreductase activity. True but uninformative; the specific activity is known. Reason: Replace with the specific RNR activity term. Proposed replacements: ribonucleoside-diphosphate reductase activity, thioredoxin disulfide as acceptor Supporting Evidence: PMID:10716984 The specific activity of the Rnr2p complexed with Rnr4p is 2,250 nmol deoxycytidine 5'-diphosphate formed per min per mg, whereas the homodimer of Rnr2p shows no activity. |
| GO:0046982 protein heterodimerization activity | IDA PMID:11526233 Structure of the yeast ribonucleotide reductase Y2Y4 heterod... | KEEP AS NON CORE | Summary: Rnr2 forms an obligate beta-beta' heterodimer with Rnr4 (crystal structure). The heterodimer is required for Rnr2 activity but the binding itself is a structural prerequisite rather than the catalytic contribution of Rnr2. Reason: Correct and informative; non-core for the radical-bearing subunit. Supporting Evidence: PMID:11526233 The heterodimer interface is similar to the mouse R2 homodimer interface in size and interacting residues PMID:10716984 the Rnr2p/Rnr4p heterodimer is the active form of the yeast RNR small subunit |
| GO:0060090 molecular adaptor activity | EXP PMID:11526233 Structure of the yeast ribonucleotide reductase Y2Y4 heterod... | MARK AS OVER ANNOTATED | Summary: Molecular adaptor activity (EXP) cited to the Y2Y4 structure paper. That paper shows Rnr2 binds iron and is the catalytic radical-bearing subunit, while the helper/scaffolding role is attributed to Y4 (Rnr4). Nothing in it shows Rnr2 bringing two other molecules together as an adaptor. Reason: Not supported by the cited full text for Rnr2; Rnr2's role is captured by RNR activity and heterodimerization. Supporting Evidence: PMID:11526233 Y4 is required for assembly of the diferric-Y* cofactor in Y2 PMID:11526233 Treatment of the crystals with Fe(II) results in difference electron density consistent with formation of a diiron center. |
| GO:0106387 'de novo' GMP biosynthetic process | RCA GO_REF:0000123 | REMOVE | Summary: YeastPathways PWY-7222-1 GO-CAM tags GDP reduction as part_of 'de novo' GMP biosynthesis. RNR makes dGDP from GDP and takes no part in GMP synthesis. Reason: Pathway-to-GO mapping error; correct process is GO:0009263, already annotated. Supporting Evidence: file:yeast/RNR2/RNR2-uniprot.txt corresponding ribonucleotides. RNR2 provides the diiron-tyrosyl radical |
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Download this section (compressed HTML)Q: Does the Rnr2-Rnr4 heterodimer stay intact through catalytic cycles in vivo, or can holo-Rnr2 act transiently without Rnr4 once the cofactor is assembled?
Q: Is nuclear Rnr2-Rnr4 merely stored, or does it contribute to dNTP synthesis near replication or repair sites?
Experiment: Compare dNTP pools and mutation rates in wtm1 and dif1 mutants versus strains forcing cytoplasmic Rnr2-Rnr4 (NES-tagged), synchronised through the cell cycle.
Hypothesis: Nuclear retention of Rnr2-Rnr4 limits dNTP synthesis outside S phase
Type: genetics / dNTP measurement
Ribonucleotide-diphosphate reductase (RNR), small subunit; the RNR complex catalyzes the rate-limiting step in dNTP synthesis and is regulated by DNA replication and DNA damage checkpoint pathways via localization of the small subunits; RNR2 has a paralog, RNR4, that arose from the whole genome duplication
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