Purification of beta-glucosidase from olive (Olea europaea L.) fruit tissue with specifically designed hydrophobic interaction chromatography and characterization of the purified enzyme

Kara, Hatibe Ertürk; Sinan, Selma; Turan, Yusuf

doi:10.1016/j.jchromb.2011.03.036

Cited by 32 publications

(40 citation statements)

References 34 publications

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“…The optimum pH and temperature of the transiently expressed enzyme activity were 5.5 and 37°C, respectively. These results coincide well with previous studies of the purified b-glucosidase enzymes from olive (Romero-Segura et al, 2009;Kara et al, 2011). Taken together, the heterologous expression of OeGLU gene in tobacco cells produced a metabolically active enzyme having similar biochemical characteristics and kinetics to the olive enzyme.…”

Section: Discussionsupporting

confidence: 91%

The C-Domain of Oleuropein β-Glucosidase Assists in Protein Folding and Sequesters the Enzyme in Nucleus

et al. 2017

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Oleuropein, a terpene-derived glycosylated secoiridoid biosynthesized exclusively by members of the Oleaceae family, is involved in a two-component defense system comprising a b-glucosidase that activates oleuropein into a toxic glutaraldehydelike structure. Oleuropein and its deglycosylated derivatives have high pharmaceutical interest. In this study we determined that the in planta heterologous expressed OeGLU, an oleuropein-specific b-glucosidase from olive (Olea europaea), had enzymatic kinetics similar to the olive native enzyme. The C terminus encompassing the nuclear localization signal sequesters the enzyme in the nucleus, and predetermines the protein-protein recognition and homodimerization. Biochemical analysis revealed that OeGLU is a homomultimer with high M r . In silico prediction modeling of the complex structure and bimolecular fluorescence complementation analyses revealed that the C terminus of OeGLU is essential for the proper assembly of an octameric form, a key conformational feature that determines the activity of the enzyme. Our results demonstrate that intrinsic characteristics of the OeGLU ensure separation from oleuropein and keep the dual-partner defensive system conditionally inactive. Upon cell destruction, the dual-partner defense system is activated and olive massively releases the arsenal of defense.

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Section: Discussionsupporting

confidence: 91%

The C-Domain of Oleuropein β-Glucosidase Assists in Protein Folding and Sequesters the Enzyme in Nucleus

et al. 2017

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“…0.5 g of acetone powder of olive fruits was added into 50 mL of buffer (4°C) with borate (pH 9.0), 5 mM EDTA, 1 mM phenylmethane sulfonyl fluoride (PMSF), and 0.25% (w/v) dithiothreitol (DTT), and solution is stirred using a Micra XRT homogenizer (Kara, Sinan, & Turan, 2011). The homogenate was centrifuged at 15,300 rpm for 30 min at 4°C (Thermo), and the solid residue after centrifugation was obtained as the raw extract (Romero-Segura, Sanz, & Perez, 2009).…”

Section: Enzyme Extractionmentioning

confidence: 99%

“…The purification of β-glucosidase from olive was achieved in two steps procedure (Kara et al, 2011). Specifically, ammonium sulfate precipitation was carried out followed by the hydrophobic interaction chromotography.…”

Section: β-Glucosidase Purificationmentioning

confidence: 99%

“…1 mL fractions of enzyme elution were obtained with the linear gradient of 1.0-0.0 M (NH 4 ) 2 SO 4 at a flow rate of 30 mL/h. Tubes with the highest enzyme activity were pooled for the further steps (Kara et al, 2011).…”

Section: β-Glucosidase Purificationmentioning

confidence: 99%

“…Well plate content was incubated at 37°C for 30 min, and the reaction was stopped by adding 70 μL of 0.5 M Na 2 CO 3 and the absorbance was measured spectrophotometrically. Enzyme activity was uttered as μmol p-nitrophenol composed per minute in the reaction medium (Kara et al, 2011;Lowry, Rosebrough, Farr, & Randall, 1951).…”

Section: Activity and Protein Measurements Of β-Glycosidasementioning

confidence: 99%

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Novel debittering process of green table olives: application ofβ-glucosidase bound onto superparamagnetic nanoparticles

Savaş

Kaya

Karaağaç

et al. 2018

CyTA - Journal of Food

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In this work, the olive β-glucosidase (β-glu) enzyme was immobilized onto superparamagnetic nanoparticles (SPMNs). Moreover, immobilized enzyme was also used for the debittering process for natural edible olives from Edremit, Turkey. Owing to SPMNs, the system can be easily removed by a simple magnet and reused many times for debittering process. The free olive β-glucosidase (E), β-glucosidase bound SPMNs (IE), free commercial β-glucosidase (CE), and commercial β-glucosidase bound SPMNs (ICE) were comparatively studied for oleuropein removal. In 6 h, the treatment of E, IE, and ICE hydrolyzed the 15.8%, 56.4%, and 80.0% of the oleuropein for 1000 g of olives, respectively, and further treatment showed that the IE and ICE reached the 100% after 22 h treatment. The results revealed that the IE and ICE with biocompatible properties of SPMNs have the economical, fast, and reusable properties for the industrial debittering process in the table olive production.Innovador método de desamargado de aceitunas verdes de mesa: aplicación de β-glucosidasa adherida a nanopartículas superparamagnéticas RESUMEN En este estudio se inmovilizó la enzima de aceituna β-glucosidasa (β-glu) en nanopartículas superparamagnéticas (SPMN); la enzima inmovilizada se utilizó en el método de desamargado de aceitunas naturales comestibles procedentes de Edremit, Turquía. Debido a las SPMN, el sistema puede ser fácilmente removido mediante un simple imán, siendo posible utilizarlo nuevamente muchas veces en el método de desamargado. Para comparar sus propiedades de eliminación de oleuropeína, se estudiaron la β-glucosidasa libre de aceituna (E), la β-glucosidasa adherida a SPMNs (IE), la β-glucosidasa libre comercial (CE) y la β-glucosidasa comercial adherida a SPMN (ICE). Después de 6 horas, los tratamientos utilizando E, IE e ICE hidrolizaron 15,8%, 56,4% y 80,0% de la oleuropeína de 1 000 g de aceitunas, respectivamente; tratamientos posteriores con IE e ICE alcanzaron 100% después de 22 horas. Estos resultados comprueban que las IE e ICE con propiedades biocompatibles de SPMN reúnen propiedades económicas y de rapidez que, aunadas a su cualidad de reusables, hace favorable su uso en el método de desamargado industrial aplicado a la producción de aceitunas de mesa. ARTICLE HISTORY

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PURIFICATION AND CHARACTERIZATION OF β‐GLUCOSIDASE FROM GREATER WAX MOTH Galleria mellonella L. (LEPIDOPTERA: PYRALIDAE)

Kara

Turan

et al. 2014

Arch Insect Biochem Physiol

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The greater wax moth, Galleria mellonella, is one of the most ruinous pests of honeycomb in the world. Beta-glucosidases are a type of digestive enzymes that hydrolytically catalyzes the beta-glycosidic linkage of glycosides. Characterization of the beta-glucosidase in G. mellonella could be a significant stage for a better comprehending of its role and establishing a safe and effective control procedure primarily against G. mellonella and also some other insect pests. Laboratory reared final instar stage larvae were randomly selected and homogenized for beta-glucosidase activity assay and subsequent analysis. The enzyme was purified to apparent homogeneity by salting out with ammonium sulfate and using sepharose-4B-l-tyrosine-1-naphthylamine hydrophobic interaction chromatography. The purification was 58-fold with an overall enzyme yield of 29%. The molecular mass of the protein was estimated as ca. 42 kDa. The purified beta-glucosidase was effectively active on para/ortho-nitrophenyl-beta-d-glucopyranosides (p-/o-NPG) with Km values of 0.37 and 1.9 mM and Vmax values of 625 and 189 U/mg, respectively. It also exhibits different levels of activity against para-nitrophenyl-β-d-fucopyranoside (p-NPF), para/ortho-nitrophenyl β-d-galactopyranosides (p-/o-NPGal) and p-nitrophenyl 1-thio-β-d-glucopyranoside. The enzyme was competitively inhibited by beta-gluconolactone and also was very tolerant to glucose against p-NPG as substrate. The Ki and IC50 values of δ-gluconolactone were determined as 0.021 and 0.08 mM while the enzyme was more tolerant to glucose inhibition with IC50 value of 213.13 mM for p-NPG.

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Purification of beta-glucosidase from olive (Olea europaea L.) fruit tissue with specifically designed hydrophobic interaction chromatography and characterization of the purified enzyme

Cited by 32 publications

References 34 publications

The C-Domain of Oleuropein β-Glucosidase Assists in Protein Folding and Sequesters the Enzyme in Nucleus

The C-Domain of Oleuropein β-Glucosidase Assists in Protein Folding and Sequesters the Enzyme in Nucleus

Novel debittering process of green table olives: application ofβ-glucosidase bound onto superparamagnetic nanoparticles

PURIFICATION AND CHARACTERIZATION OF β‐GLUCOSIDASE FROM GREATER WAX MOTH Galleria mellonella L. (LEPIDOPTERA: PYRALIDAE)

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