BGLU15 Antibody
Description
BGLU15: Enzyme Overview
BGLU15 (At2g44450) is a glycoside hydrolase family 1 enzyme in Arabidopsis thaliana that catalyzes the hydrolysis of flavonol 3-O-β-glucoside-7-O-α-rhamnosides during recovery from nitrogen-deprivation/low-temperature (NDLT) stress . Key characteristics include:
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Substrate Specificity: Preferentially cleaves flavonol bisglycosides (e.g., kaempferol 3-O-β-glucoside-7-O-α-rhamnoside [K3G7R] and quercetin 3-O-β-glucoside-7-O-α-rhamnoside [Q3G7R]) over monoglucosides or other conjugates .
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Kinetic Parameters: Demonstrates a k<sub>cat</sub>/K<sub>m</sub> of 2.1–4.3 × 10<sup>4</sup> M<sup>−1</sup>s<sup>−1</sup> for flavonol bisglycosides, with a lower efficiency for monoglucosides (e.g., k<sub>cat</sub>/K<sub>m</sub> = 1.3 × 10<sup>4</sup> M<sup>−1</sup>s<sup>−1</sup> for quercetin 3-O-β-glucoside) .
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Localization: Loosely associated with the cell wall, necessitating flavonol bisglycoside efflux from vacuoles for enzymatic activity .
Gene Expression and Stress Response
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Transcript Induction: BGLU15 mRNA levels spike 280–330% within 1 day of NDLT recovery in shoots, coinciding with flavonol bisglycoside catabolism .
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Mutant Studies: bglu15 T-DNA insertional mutants (bglu15-1 and bglu15-2) fail to degrade flavonol 3-O-β-glucoside-7-O-α-rhamnosides during stress recovery, confirming its essential role .
Biochemical Characterization
Recombinant BGLU15 (72.6 kDa, thioredoxin-His<sub>6</sub>-tagged) was expressed in Escherichia coli and purified via immobilized metal affinity chromatography (IMAC) . Key enzymatic properties:
| Substrate | K<sub>m</sub> (µM) | k<sub>cat</sub> (s<sup>−1</sup>) | k<sub>cat</sub>/K<sub>m</sub> (M<sup>−1</sup>s<sup>−1</sup>) |
|---|---|---|---|
| K3G7R | 36 ± 4 | 1.5 ± 0.1 | 4.3 × 10<sup>4</sup> |
| Q3G7R | 60 ± 7 | 1.7 ± 0.2 | 2.1 × 10<sup>4</sup> |
| Quercetin 3-O-β-glucoside | 110 ± 12 | 1.4 ± 0.1 | 1.3 × 10<sup>4</sup> |
Data derived from discontinuous in vitro assays using 500 µM substrate concentrations .
Implications for Antibody Development
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Antibody Generation: Polyclonal or monoclonal antibodies targeting BGLU15’s unique epitopes for tissue-specific localization studies.
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Functional Assays: Using antibodies to quantify BGLU15 protein levels under stress conditions or in mutant backgrounds.
Limitations and Knowledge Gaps
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- BGLU15 exhibits a preference for hydrolyzing the 3-O-beta-glucosides of flavonols. However, it does not cleave quercetin 3-O-alpha-rhamnoside, quercetin 3-O-beta-galactoside, or rutin. PMID: 25468534
