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Literature summary for 4.2.3.103 extracted from
- A pair of residues that interactively affect diterpene synthase product outcome (2017), ACS Chem. Biol., 12, 862-867 .
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| gene KSL5i, sequence comparisons and phylogenetic analysis and tree, recombinant expression of His-tagged wild-type and mutant enzymes without the N-terminal transit peptide sequence in Escherichia coli | Oryza sativa Indica Group |
| gene KSL6, sequence comparisons and phylogenetic analysis and tree, recombinant expression of His-tagged wild-type and mutant enzymes without the N-terminal transit peptide sequence in Escherichia coli | Oryza sativa Japonica Group |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| I664T | site-directed mutagenesis, the OsKSL5i:I664T mutant produces only ent-pimara-8(14),15-diene | Oryza sativa Indica Group |
| additional information | of the three differences in active site residues between the orthologous OsKSL5j and OsKSL5i, switching the Ile at position 664 in OsKSL5i to the Thr found in OsKSL5j is sufficient to short circuit the production of ent-isokaurene, as OsKSL5i:I664T specifically produces ent-pimara-8(14),15-diene instead. But the reciprocal residue switch leads to production of a mixture of tetracycles ent-isokaurene, ent-kaurene, and ent-atiserene (i.e., by OsKSL5j:T664I), rather than the almost exclusive production of ent-isokaurene observed with OsKSL5i | Oryza sativa Indica Group |
| V661L | site-directed mutagenesis, substitution of Leu for the Val found at this secondary position only slightly decreases catalytic fidelity, the mutant still predominantly produces ent-isokaurene, with only slight increases in the relative amounts of ent-kaur-16-ene and ent-atiserene | Oryza sativa Japonica Group |
| V661L | site-directed mutagenesis, the mutant single residue switch, OsKSL5i:V661L, leads to reduced catalytic specificity and production of a mixture of ent-isokaurene, ent-keurene, and ent-atiserene, closely resembling that mediated by the OsKSL5j:T664I mutant (EC 4.2.3.30), although it does not affect the specific production of ent-pimara-8(14),15-diene | Oryza sativa Indica Group |
Localization
| Localization | Comment | Organism | GeneOntology No. | Textmining |
|---|---|---|---|---|
| chloroplast | the enzyme contains a N-terminal transit peptide sequence | Oryza sativa Japonica Group | 9507 | - |
| chloroplast | the enzyme contains a N-terminal transit peptide sequence | Oryza sativa Indica Group | 9507 | - |
Metals/Ions
| Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|
| Mg2+ | required | Oryza sativa Japonica Group |  |
| Mg2+ | required | Oryza sativa Indica Group | |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| ent-copalyl diphosphate | Oryza sativa Japonica Group | - |
ent-isokaurene + diphosphate | - |
? | |
| ent-copalyl diphosphate | Oryza sativa Indica Group | - |
ent-isokaur-15-ene + diphosphate | - |
? | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Oryza sativa Indica Group | A4KAG7 | - |
- |
| Oryza sativa Japonica Group | A4KAG8 | - |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant His-tagged wild-type and mutant enzymes without the N-terminal transit peptide sequence from Escherichia coli by nickel affinity chromatography | Oryza sativa Japonica Group |
| recombinant His-tagged wild-type and mutant enzymes without the N-terminal transit peptide sequence from Escherichia coli by nickel affinity chromatography | Oryza sativa Indica Group |
Source Tissue
| Source Tissue | Comment | Organism | Textmining |
|---|---|---|---|
| leaf | - |
Oryza sativa Japonica Group | - |
| leaf | - |
Oryza sativa Indica Group | - |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| ent-copalyl diphosphate | - |
Oryza sativa Japonica Group | ent-isokaurene + diphosphate | - |
? | |
| ent-copalyl diphosphate | - |
Oryza sativa Indica Group | ent-isokaur-15-ene + diphosphate | - |
? | |
| additional information | GC-MS reaction product analysis. The distinct function of two alleles of a KSL from rice, OsKSL5i and OsKSLj, identifies a single residue switch with a profound effect on not only OsKSL5 product outcome but also that of land plant KSs more broadly, specifically, replacement of a key isoleucine with threonine, which interrupts formation of the tetracyclic ent-isokaurene at the tricyclic stage, leading to production of ent-pimaradiene instead | Oryza sativa Indica Group | ? | - |
? | |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| kaurene synthase-like enzyme 5 | - |
Oryza sativa Indica Group |
| kaurene synthase-like enzyme 6 | - |
Oryza sativa Japonica Group |
| KS-like 5i | - |
Oryza sativa Indica Group |
| KS-like 6 | - |
Oryza sativa Japonica Group |
| OsKSL5i | - |
Oryza sativa Indica Group |
| OsKSL6 | - |
Oryza sativa Japonica Group |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 37 | - |
assay at | Oryza sativa Japonica Group |
| 37 | - |
assay at | Oryza sativa Indica Group |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 7 | - |
assay at | Oryza sativa Japonica Group |
| 7 | - |
assay at | Oryza sativa Indica Group |
General Information
| General Information | Comment | Organism |
|---|---|---|
| evolution | in the plant kingdom, where the labdane-related diterpenoids (LRDs) are predominantly found, the class I and II diterpene cyclases seem to have most directly evolved via gene duplication and neofunctionalization of the ent-kaurene synthases (KSs) required in all vascular plants for gibberellin hormone biosynthesis. Accordingly, these enzymes are often termed KS-like (KSL) and form a distinct subfamily within the plant terpene synthase family. Like class I terpene synthases more generally, the KS(L)s carry out catalysis in a highly conserved alpha-helical bundle domain that contains two signature motifs. These DDxxD and NDxx(S/T)xxxE sequences are involved in ligation of a trinuclear Mg2+ cluster to promote substrate binding and the subsequent initiating ionization of the allylic diphosphate ester bond | Oryza sativa Japonica Group |
| evolution | in the plant kingdom, where the labdane-related diterpenoids (LRDs) are predominantly found, the class I and II diterpene cyclases seem to have most directly evolved via gene duplication and neofunctionalization of the ent-kaurene synthases (KSs) required in all vascular plants for gibberellin hormone biosynthesis. Accordingly, these enzymes are often termed KS-like (KSL) and form a distinct subfamily within the plant terpene synthase family. Like class I terpene synthases more generally, the KS(L)s carry out catalysis in a highly conserved alpha-helical bundle domain that contains two signature motifs. These DDxxD and NDxx(S/T)xxxE sequences are involved in ligation of a trinuclear Mg2+ cluster to promote substrate binding and the subsequent initiating ionization of the allylic diphosphate ester bond. OsKSL5 belongs to an early diverging KSL lineage within monocots and is relatively distantly related to any KSs | Oryza sativa Indica Group |
| additional information | conserved residues at positions 661 (secondary) and 664 (primary) are responsible for substrate specificity. Two functionally distinct alleles of Oryza sativa OsKSL5, which react with ent-copalyl diphosphate (ent-copalyl diphosphate), with that from subspecies japonica (OsKSL5j, EC 4.2.3.30) producing ent-pimara-8(14),15-diene and that from subspecies indica (OsKSL5i, EC 4.2.3.103) producing ent-(iso)kaur-15-ene instead | Oryza sativa Indica Group |
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Release 2023.1 (January 2023)
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