Abstract
β-Glucosidases (EC 3.2.1.21) split β-glucosidic linkages at the non-reducing end of glucosides and oligosaccharides to release β-D-glucose. One of the important functions of plant β-glucosidase is deglucosylation of inactive glucosides of phytohormones to regulate levels of active hormones. Tuberonic acid is a jasmonate-related compound that shows tuber-inducing activity in the potato. We have identified two enzymes, OsTAGG1 and OsTAGG2, that have hydrolytic activity towards tuberonic acid β-D-glucoside in rice (Oryza sativa L.). The expression of OsTAGG2 is upregulated by wounding and by methyl jasmonate, suggesting that this isozyme is involved in responses to biotic stresses and wounding, but the physiological substrate of OsTAGG2 remains ambiguous. In this study, we produced recombinant OsTAGG2 in Pichia pastoris (rOsTAGG2P), and investigated its substrate specificity in detail. From 1 L of culture medium, 2.1 mg of purified recombinant enzyme was obtained by ammonium sulfate precipitation and Ni-chelating column chromatography. The specific activity of rOsTAGG2P (182 U/mg) was close to that of the native enzyme (171 U/mg), unlike recombinant OsTAGG2 produced in Escherichia coli, which had approximately 3-fold lower specific activity than the native enzyme. The optimum pH and temperature for rOsTAGG2P were pH 3.4 and 60 °C. After pH and heat treatments, the enzyme retained its original activity in a pH range of 3.4-9.8 and below 55 °C. Native OsTAGG2 and rOsTAGG2P showed 4.5-4.7-fold higher activities towards salicylic acid β-D-glucoside, an inactive storage-form of salicylic acid, than towards tuberonic acid β-D-glucoside (TAG), although OsTAGG2 was originally isolated from rice based on TAG-hydrolytic activity.
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Claims 8
Claim · model-written summary
Recombinant OsTAGG2 yield was 2.1 mg per litre of culture medium.
- Taxon
- Rice
Oryza sativat0 - Outcome
- purified recombinant enzymee0
- Value
- 2.1 mge0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“From 1 L of culture medium, 2.1 mg of purified recombinant enzyme was obtained”
t0“Rice”
Claim · model-written summary
Recombinant enzyme specific activity was 182 U/mg, near native enzyme at 171 U/mg.
- Taxon
- RiceOryza sativat0
- Gene
- rOsTAGG2Pe0
- Outcome
- specific activitye0
- Value
- 182 U/mge0
- Qualifier
- 171 U/mge0
- Comparator
- native enzymee0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“The specific activity of rOsTAGG2P (182 U/mg) was close to that of the native enzyme (171 U/mg)”
t0“Rice”
Claim · model-written summary
E. coli recombinant OsTAGG2 had about threefold lower specific activity than native enzyme.
- Taxon
- RiceOryza sativat0
- Gene
- OsTAGG2e0
- Outcome
- specific activitye0
- Direction
- lowere0
- Value
- approximately 3-folde0
- Comparator
- native enzymee0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“recombinant OsTAGG2 produced in Escherichia coli, which had approximately 3-fold lower specific activity than the native enzyme.”
t0“Rice”
Claim · model-written summary
Recombinant OsTAGG2 optimum pH was 3.4.
- Taxon
- RiceOryza sativat0
- Gene
- rOsTAGG2Pe0
- Outcome
- optimum pHe0
- Value
- pH 3.4e0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“The optimum pH and temperature for rOsTAGG2P were pH 3.4 and 60 °C.”
t0“Rice”
Claim · model-written summary
Recombinant OsTAGG2 optimum temperature was 60 °C.
- Taxon
- RiceOryza sativat0
- Gene
- rOsTAGG2Pe0
- Outcome
- temperaturee0
- Value
- 60 °Ce0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“The optimum pH and temperature for rOsTAGG2P were pH 3.4 and 60 °C.”
t0“Rice”
Claim · model-written summary
Enzyme activity persisted after treatments across pH 3.4–9.8.
- Taxon
- RiceOryza sativat0
- Outcome
- original activitye0
- Value
- 3.4-9.8e0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“the enzyme retained its original activity in a pH range of 3.4-9.8 and below 55 °C.”
t0“Rice”
Claim · model-written summary
Enzyme activity persisted after heat treatment below 55 °C.
- Taxon
- RiceOryza sativat0
- Outcome
- original activitye0
- Value
- 55 °Ce0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“the enzyme retained its original activity in a pH range of 3.4-9.8 and below 55 °C.”
t0“Rice”
Claim · model-written summary
Native and recombinant OsTAGG2 showed 4.5–4.7-fold higher activity on salicylic acid glucoside than TAG.
- Taxon
- RiceOryza sativat0
- Gene
- OsTAGG2 and rOsTAGG2Pe0
- Outcome
- activities towards salicylic acid β-D-glucosidee0
- Direction
- highere0
- Value
- 4.5-4.7-folde0
- Comparator
- tuberonic acid β-D-glucoside (TAG)e0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Native OsTAGG2 and rOsTAGG2P showed 4.5-4.7-fold higher activities towards salicylic acid β-D-glucoside, an inactive storage-form of salicylic acid, than towards tuberonic acid β-D-glucoside (TAG)”
t0“Rice”