GA2 (ent-kaurene synthase, KS; At1g79460) is a plastid-targeted class I diterpene synthase of the terpene synthase family that converts ent-copalyl diphosphate, the product of GA1/CPS, into the tetracyclic hydrocarbon ent-kaur-16-ene with release of diphosphate, the second committed reaction of gibberellin biosynthesis. Catalysis uses Mg2+ bound by DDXXD-type motifs to ionize the diphosphate. The protein is imported into chloroplasts/plastids, where it is a soluble stromal enzyme, and is expressed throughout the plant. ga2 mutants (e.g. ga2-1, carrying a premature stop that removes the C-terminal catalytic region) are gibberellin-deficient dwarfs. A single active-site residue change converts land-plant KS enzymes into pimaradiene synthases, illustrating how specialized diterpene metabolism derives from this enzyme.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000287 magnesium ion binding | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Mg2+ binding by the class I terpene synthase domain (DDXXD/NSE-DTE motifs coordinate three Mg2+ ions that trigger diphosphate ionization). Reason: Mg2+ is the catalytic cofactor of KS; cofactor binding is retained as non-core supporting the ent-kaurene synthase activity. Supporting Evidence: PMID:21602811 Class I cyclases contain DDXXD and (N,D)DXX(S,T)XXXE motifs; boldface residues generally coordinate to three Mg2+ ions, which trigger the ionization of an isoprenoid diphosphate group to generate a reactive carbocation intermediate. |
| GO:0000287 magnesium ion binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: Mg2+ binding from the InterPro terpene synthase metal-binding domain. Reason: Correct cofactor binding for a class I diterpene synthase; non-core. Supporting Evidence: PMID:21602811 Class I cyclases contain DDXXD and (N,D)DXX(S,T)XXXE motifs; boldface residues generally coordinate to three Mg2+ ions, which trigger the ionization of an isoprenoid diphosphate group to generate a reactive carbocation intermediate. |
| GO:0009507 chloroplast | HDA PMID:18431481 Sorting signals, N-terminal modifications and abundance of t... | ACCEPT | Summary: Detected in the chloroplast proteome by mass spectrometry. Reason: Consistent with GFP targeting and import into isolated chloroplasts. Supporting Evidence: PMID:11722763 The AtKS1-GFP fusion is also imported by isolated pea chloroplasts. |
| GO:0009507 chloroplast | IBA GO_REF:0000033 | ACCEPT | Summary: Chloroplast localization inherited within the plant KS clade. Reason: Consistent with experimental data. Supporting Evidence: PMID:11722763 We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. |
| GO:0009507 chloroplast | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular location mapping (chloroplast). Reason: Consistent with experimental data. Supporting Evidence: PMID:11722763 We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. |
| GO:0009507 chloroplast | ISM GO_REF:0000122 | ACCEPT | Summary: AtSubP prediction (chloroplast). Reason: Consistent with experimental data. Supporting Evidence: PMID:11722763 We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. |
| GO:0009507 chloroplast | TAS PMID:9536043 The GA2 locus of Arabidopsis thaliana encodes ent-kaurene sy... | ACCEPT | Summary: Author statement of chloroplast localization in the GA2 cloning paper. Reason: Later confirmed experimentally by GFP fusion and import assays. Supporting Evidence: PMID:11722763 The AtKS1-GFP fusion is also imported by isolated pea chloroplasts. |
| GO:0009536 plastid | TAS PMID:9536043 The GA2 locus of Arabidopsis thaliana encodes ent-kaurene sy... | ACCEPT | Summary: Plastid localization (TAS). Reason: Correct; KS acts in plastids in all tissues, of which the chloroplast is the leaf form. Supporting Evidence: PMID:11722763 We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. |
| GO:0009570 chloroplast stroma | IDA PMID:11722763 A plastid envelope location of Arabidopsis ent-kaurene oxida... | ACCEPT | Summary: AtKS1-GFP is targeted to chloroplasts and imported by isolated pea chloroplasts; KS is a soluble stromal enzyme lacking membrane anchors. Reason: Best-supported specific location for KS; used as the core-function location. Supporting Evidence: PMID:11722763 We show that the N-terminal pre-sequences of the Arabidopsis thaliana terpene cyclases copalyl diphosphate synthase (AtCPS1) and ent-kaurene synthase (AtKS1) direct GFP to chloroplasts in transient assays following microprojectile bombardment of tobacco leaves. PMID:11722763 The AtKS1-GFP fusion is also imported by isolated pea chloroplasts. |
| GO:0009686 gibberellin biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: KS converts ent-CPP to ent-kaurene, the second plastid step of GA biosynthesis; ga2 mutants are GA-deficient dwarfs. Reason: Conserved function of the land-plant KS clade. Supporting Evidence: PMID:17456599 conversion of the ent-kaurene synthases found in all higher plants for gibberellin phytohormone biosynthesis to the production of pimaradiene |
| GO:0009686 gibberellin biosynthetic process | IDA PMID:9536043 The GA2 locus of Arabidopsis thaliana encodes ent-kaurene sy... | ACCEPT | Summary: KS converts ent-CPP to ent-kaurene, the second plastid step of GA biosynthesis; ga2 mutants are GA-deficient dwarfs. Reason: Recombinant AtKS converts CPP to ent-kaurene. Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. PMID:9536043 The ga2 mutant of Arabidopsis thaliana is a gibberellin-deficient dwarf. |
| GO:0009686 gibberellin biosynthetic process | IEA GO_REF:0000041 | ACCEPT | Summary: KS converts ent-CPP to ent-kaurene, the second plastid step of GA biosynthesis; ga2 mutants are GA-deficient dwarfs. Reason: UniPathway mapping consistent with experimental data. Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. |
| GO:0009740 gibberellic acid mediated signaling pathway | TAS PMID:9536043 The GA2 locus of Arabidopsis thaliana encodes ent-kaurene sy... | MARK AS OVER ANNOTATED | Summary: KS makes a hormone precursor; it is not part of GA signal transduction. Reason: Biosynthesis of the ligand is upstream of, not part of, the GA-GID1-DELLA signaling pathway; the process role is captured by gibberellin biosynthetic process. Supporting Evidence: PMID:9536043 The ga2 mutant of Arabidopsis thaliana is a gibberellin-deficient dwarf. |
| GO:0009899 ent-kaurene synthase activity | EXP PMID:17456599 Following evolution's lead to a single residue switch for di... | ACCEPT | Summary: Xu et al. 2007 assayed land-plant ent-kaurene synthases, including AtKS, and identified an Ile-to-Thr switch that redirects product outcome to pimaradiene. Reason: Experimental characterization of AtKS ent-kaurene synthase activity. Supporting Evidence: PMID:17456599 conversion of the ent-kaurene synthases found in all higher plants for gibberellin phytohormone biosynthesis to the production of pimaradiene |
| GO:0009899 ent-kaurene synthase activity | IDA PMID:9536043 The GA2 locus of Arabidopsis thaliana encodes ent-kaurene sy... | ACCEPT | Summary: Recombinant AtKS fusion protein converted copalyl diphosphate to ent-kaurene; ga2-1 truncated protein is inactive. Reason: Core activity (RHEA:22220). Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. PMID:9536043 The recombinant AtKS protein derived from the ga2-1 mutant is truncated by 14 kD at the C-terminal end and does not contain significant KS activity in vitro. |
| GO:0009899 ent-kaurene synthase activity | IEA GO_REF:0000120 | ACCEPT | Summary: EC/Rhea-based mapping. Reason: Matches experimental evidence. Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. |
| GO:0010333 terpene synthase activity | IBA GO_REF:0000033 | MODIFY | Summary: Terpene synthase activity propagated from a PANTHER node shared by CPS and KS. Reason: Correct for KS (ent-kaurene synthase activity is_a terpene synthase activity) but less specific than the experimentally established activity. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN001272394 SUPPORTS TRANSFER Node spans CPS and KS; terpene synthase activity is correct for KS but the specific ent-kaurene synthase term is available and experimentally supported on the target. AGI_LocusCode:AT1G79460 SUPPORTS TRANSFER GA2/KS itself has direct experimental ent-kaurene synthase evidence (PMID:9536043). Proposed replacements: ent-kaurene synthase activity Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. |
| GO:0010333 terpene synthase activity | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO terpene synthase activity. Reason: Correct parent; use the specific activity. Proposed replacements: ent-kaurene synthase activity Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. |
| GO:0016102 diterpenoid biosynthetic process | IEA GO_REF:0000002 | MODIFY | Summary: Diterpenoid biosynthetic process (InterPro2GO). Reason: Correct but general; ent-kaurene is the GA precursor. Proposed replacements: gibberellin biosynthetic process Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. |
| GO:0016114 terpenoid biosynthetic process | IEA GO_REF:0000120 | MODIFY | Summary: Terpenoid biosynthetic process. Reason: Correct but general. Proposed replacements: gibberellin biosynthetic process Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. |
| GO:0016829 lyase activity | IEA GO_REF:0000002 | MODIFY | Summary: Lyase activity (InterPro2GO). Reason: KS is a diphosphate-eliminating class I cyclase (EC 4.2.3.19), so lyase activity is correct but uninformative. Proposed replacements: ent-kaurene synthase activity Supporting Evidence: PMID:9536043 Taken together, our results show that the GA2 locus encodes KS. |
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Download this section (compressed HTML)Q: Do CPS and KS physically associate in the plastid stroma to channel ent-CPP, and does such association affect flux into gibberellin versus other diterpenoids?
Experiment: Test CPS-KS proximity in planta (e.g. BiFC or proximity labelling in plastids) and measure ent-CPP leakage in lines with altered CPS:KS stoichiometry.
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