EC Number | Cloned (Comment) | Organism |
---|---|---|
4.2.3.15 | mutated enzymes are generated using the QuickChange II site-directed mutagenesis kit. The PCR-based mutagenesis protocol is performed using pET200-D-TOPO harbouring the respective AcTPS2 cDNAs as template. The single-site mutant enzymes overexpressed in Escherichia coli. | Actinidia chinensis |
4.2.3.46 | mutated enzymes are generated using the QuickChange II site-directed mutagenesis kit. The PCR-based mutagenesis protocol is performed using pET-30a harbouring the MdAFS1 cDNAs as template. The single-site mutant enzymes overexpressed in Escherichia coli. | Malus domestica |
EC Number | Protein Variants | Comment | Organism |
---|---|---|---|
4.2.3.15 | K514A | AcTPS2 mutant, 90% reduction in monoterpene synthase activity compared to wild-type enzyme | Actinidia chinensis |
4.2.3.15 | K514S | AcTPS2 mutant, presence of Ser is not able to rescue any of the activity lost after the removal of the Lys residue | Actinidia chinensis |
4.2.3.46 | D484A | 85% loss of sesquiterpene synthase activity compared with wild-type enzyme when K+ is present | Malus domestica |
4.2.3.46 | D484A | 85% loss of sesquiterpene synthase activity compared with wild-type enzyme. Little change in K+-independent activity | Malus domestica |
4.2.3.46 | S485A | exhibits marginally increased sesquiterpene synthase activities, and an approximate 2fold increase in monoterpene synthase activity compared with the wild-type enzyme | Malus domestica |
4.2.3.46 | S485A | mutant exhibits marginally increased sesquiterpene synthase activities, and an approximate 2fold increase in monoterpene synthase activity compared with the wild-type enzyme | Malus domestica |
4.2.3.46 | S487A | 95% decrease in sesquiterpene synthase activity compared with wild-type enzyme when K+ is present | Malus domestica |
4.2.3.46 | S487A | 95% decrease in sesquiterpene synthase activity compared with wild-type enzyme. Little change in K+-independent activity | Malus domestica |
4.2.3.46 | S487K | The S487K mutant has 35-45% of the wild-type activity with farnesyl diphosphate, with no significant alterations in the alpha-farnesene isomer ratios produced and a small decrease in catalytic efficiency | Malus domestica |
4.2.3.46 | S487K | mutant has 3545% of the wild-type activity with farnesyl diphosphate, with no significant alterations in the alpha-farnesene isomer ratios produced and a small decrease in catalytic efficiency | Malus domestica |
4.2.3.46 | S488A | mutant shows decreases in both sesqui- and monoterpene synthase activities compared with the WT enzyme, with mono-TPS activity being reduced more than sesquiterpene synthase activity. Sesquiterpene synthase (alpha-farnesene) products produced by the mutated and wild-type enzymes are identical, there are no significant alterations in the ratios of the alpha-farnesene isomers produced. | Malus domestica |
4.2.3.46 | S488A | mutant shows decreases in both sesqui- and mono-terpene synthases activities, compared with the wild-type enzyme, with mono-terpene synthases activity being reduced more than sesqui-terpene synthases activity | Malus domestica |
EC Number | Metals/Ions | Comment | Organism | Structure |
---|---|---|---|---|
4.2.3.15 | Mg2+ | 10 mM are included in farnesyl diphosphate activity assay | Actinidia chinensis | |
4.2.3.15 | additional information | wild-type AcTPS2 is significantly more active when KCl is absent from the activity buffer | Actinidia chinensis | |
4.2.3.46 | K+ | MdAFS1 retains up to 12% of its activity in the absence of K+, enzyme contains K+ binding region, MdAFS1 exhibits a type II K+ response, MdAFS1 is not absolutely dependent upon M+ its unequivocal classification as type I or type II K+ activated, or that of any other terpene synthases, will not be possibl, then type I enzymes can exhibit type II kinetics and vice versa. | Malus domestica | |
4.2.3.46 | K+ | activity is dependent on K+, the potassium binding region is defined | Malus domestica | |
4.2.3.46 | Mg2+ | 10 mM are included in farnesyl diphosphate activity assay | Malus domestica |
EC Number | Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|---|
4.2.3.15 | additional information | Actinidia chinensis | terpenes produced from the wild-type and mutant enzymes are identical, showing low levels of beta-myrcene from AcTPS2-K514A | ? | - |
? | |
4.2.3.46 | (2E,6E)-farnesyl diphosphate | Malus domestica | commentary | (3E,6E)-alpha-farnesene + diphosphate | - |
? | |
4.2.3.46 | additional information | Malus domestica | the monoterpene synthase ((E)-beta-ocimene and beta-myrcene) and sesquiterpene synthase (alpha-farnesene) products produced by the mutated and wild-type enzymes are identical, there are no significant alterations in the ratios of the alpha-farnesene isomers produced | ? | - |
? |
EC Number | Organism | UniProt | Comment | Textmining |
---|---|---|---|---|
4.2.3.15 | Actinidia chinensis | - |
- |
- |
4.2.3.46 | Malus domestica | B2ZZ11 | - |
- |
4.2.3.46 | Malus domestica | Q84LB2 | - |
- |
EC Number | Purification (Comment) | Organism |
---|---|---|
4.2.3.15 | recombinant AcTPS2 protein is extracted and purified | Actinidia chinensis |
4.2.3.46 | recombinant MdAFS1protein is extracted and purified | Malus domestica |
EC Number | Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|---|
4.2.3.46 | additional information | - |
S487K mutant has 35-45% of the wild-type activity with farnesyl diphosphate with no significant alterations in the alpha-farnesene isomer ratios produced and a small decrease in catalytic efficiency (wild-type and S487K respective kcat/Km values of 17.5 and 13.3 mM/s) | Malus domestica |
EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|---|
4.2.3.15 | additional information | terpenes produced from the wild-type and mutant enzymes are identical, showing low levels of beta-myrcene from AcTPS2-K514A | Actinidia chinensis | ? | - |
? | |
4.2.3.46 | (2E,6E)-farnesyl diphosphate | - |
Malus domestica | (3E,6E)-alpha-farnesene + diphosphate | - |
? | |
4.2.3.46 | (2E,6E)-farnesyl diphosphate | commentary | Malus domestica | (3E,6E)-alpha-farnesene + diphosphate | - |
? | |
4.2.3.46 | geranyl diphosphate | poor substrate | Malus domestica | (E)-beta-ocimene + beta-myrcene | - |
? | |
4.2.3.46 | additional information | the monoterpene synthase ((E)-beta-ocimene and beta-myrcene) and sesquiterpene synthase (alpha-farnesene) products produced by the mutated and wild-type enzymes are identical, there are no significant alterations in the ratios of the alpha-farnesene isomers produced | Malus domestica | ? | - |
? |
EC Number | Synonyms | Comment | Organism |
---|---|---|---|
4.2.3.15 | AcTPS2 | - |
Actinidia chinensis |
4.2.3.15 | myrcene synthase | - |
Actinidia chinensis |
4.2.3.46 | alpha-farnesene synthase | - |
Malus domestica |
4.2.3.46 | MdAFS1 | - |
Malus domestica |
EC Number | Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|---|
4.2.3.15 | 30 | - |
farnesyl diphosphate and geranyl diphosphate activity assay | Actinidia chinensis |
4.2.3.46 | 30 | - |
farnesyl diphosphate and geranyl diphosphate activity assay | Malus domestica |
EC Number | Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|---|
4.2.3.46 | 0.000005 | - |
geranyl diphosphate | mutant S487A | Malus domestica | |
4.2.3.46 | 0.000005 | - |
geranyl diphosphate | pH 7.5, 30°C, mutant enzyme S487A | Malus domestica | |
4.2.3.46 | 0.000034 | - |
geranyl diphosphate | mutant D484A | Malus domestica | |
4.2.3.46 | 0.000034 | - |
geranyl diphosphate | pH 7.5, 30°C, mutant enzyme D484A | Malus domestica | |
4.2.3.46 | 0.000052 | - |
geranyl diphosphate | mutant S488A | Malus domestica | |
4.2.3.46 | 0.000052 | - |
geranyl diphosphate | pH 7.5, 30°C, mutant enzyme S488A | Malus domestica | |
4.2.3.46 | 0.000281 | - |
geranyl diphosphate | wild-type | Malus domestica | |
4.2.3.46 | 0.000281 | - |
geranyl diphosphate | pH 7.5, 30°C, wild-type enzyme | Malus domestica | |
4.2.3.46 | 0.000588 | - |
geranyl diphosphate | mutant S485A | Malus domestica | |
4.2.3.46 | 0.000588 | - |
geranyl diphosphate | pH 7.5, 30°C, mutant enzyme S485A | Malus domestica | |
4.2.3.46 | 0.0026 | - |
farnesyl diphosphate | mutant S487A, 50 mM KCl, 10 mM MgCl2 | Malus domestica | |
4.2.3.46 | 0.0026 | - |
(2E,6E)-farnesyl diphosphate | pH 7.5, 30°C, mutant enzyme S487A | Malus domestica | |
4.2.3.46 | 0.0062 | - |
farnesyl diphosphate | without K+ | Malus domestica | |
4.2.3.46 | 0.0086 | - |
farnesyl diphosphate | mutant D484A, 50 mM KCl, 10 mM MgCl2 | Malus domestica | |
4.2.3.46 | 0.0086 | - |
(2E,6E)-farnesyl diphosphate | pH 7.5, 30°C, mutant enzyme D484A | Malus domestica | |
4.2.3.46 | 0.022 | - |
farnesyl diphosphate | without K+, sesquiterpene synthase activity in mutant S487K is independent of K+ | Malus domestica | |
4.2.3.46 | 0.039 | - |
farnesyl diphosphate | mutant S488A, 50 mM KCl, 10 mM MgCl2 | Malus domestica | |
4.2.3.46 | 0.039 | - |
(2E,6E)-farnesyl diphosphate | pH 7.5, 30°C, mutant enzyme S488A | Malus domestica | |
4.2.3.46 | 0.0533 | - |
farnesyl diphosphate | wild-type MdAFS, 50 mM KCl, 10 mM MgCl2 | Malus domestica | |
4.2.3.46 | 0.0553 | - |
(2E,6E)-farnesyl diphosphate | pH 7.5, 30°C, wild-type enzyme | Malus domestica | |
4.2.3.46 | 0.0613 | - |
farnesyl diphosphate | mutant S485A, 50 mM KCl, 10 mM MgCl2 | Malus domestica | |
4.2.3.46 | 0.0613 | - |
(2E,6E)-farnesyl diphosphate | pH 7.5, 30°C, mutant enzyme S485A | Malus domestica |
EC Number | pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|---|
4.2.3.15 | 7.5 | - |
farnesyl diphosphate activity assay | Actinidia chinensis |
4.2.3.46 | 7.5 | - |
farnesyl diphosphate activity assay | Malus domestica |