ß-glucosidases in the rice weevil, Sitophilus oryzae:Purification, properties, and activity levels in wheat- and legume-feeding strains

Baker, James E.; Woo, S.M.

doi:10.1016/0965-1748(92)90146-6

Cited by 38 publications

(21 citation statements)

References 18 publications

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“…However, the second bglucosidase displayed a much lower affinity towards NPbGlc, which is quite unusual for insects. Only once before, in a coleopteran species, Sitophilus oryzae (Baker and Woo, 1992), was the existence of a b-glucosidase with such a low Michaelis constant reported.…”

Section: Kinetic Properties Of B-glucosidase Activitymentioning

confidence: 94%

The interplay between toxin-releasing β-glucosidase and plant iridoid glycosides impairs larval development in a generalist caterpillar, Grammia incorrupta (Arctiidae)

Pankoke¹,

Bowers

Dobler³

2012

Insect Biochemistry and Molecular Biology

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Section: Kinetic Properties Of B-glucosidase Activitymentioning

confidence: 94%

The interplay between toxin-releasing β-glucosidase and plant iridoid glycosides impairs larval development in a generalist caterpillar, Grammia incorrupta (Arctiidae)

Pankoke¹,

Bowers

Dobler³

2012

Insect Biochemistry and Molecular Biology

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“…The fusion peptide (CBP) was not removed from the recombinant Sfβgly50 used in the assays. The hydrolysis of NPβglycosides was followed by the NP release [14] and MUβGlc hydrolysis by MU fluorescence [17]. The kinetic parameters ( k cat and K m ) were determined employing 10 different substrate concentrations (bracketing the K m values) and the data were fitted to Michaelis–Menten equation using the enzfitter software (Elsevier‐Biosoft, Cambridge, UK).…”

Section: Methodsmentioning

confidence: 99%

The role of amino‐acid residues Q39 and E451 in the determination of substrate specificity of the Spodoptera frugiperdaβ‐glycosidase

Marana

Terra

Ferreira

2002

European Journal of Biochemistry

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The design of b-glycosidases with planed substrate specificity for biotechnological application has received little attention. This is mostly a consequence of the lack of data on the molecular basis of the b-glycosidase specificity, namely data on the energy of the noncovalent interactions in the enzymetransition state complex. In an attempt to fill this gap, sitedirected mutagenesis and enzyme steady-state kinetic experiments with different substrates were conducted, using as model a digestive b-glycosidase (glycoside hydrolase family 1) from Spodoptera frugiperda (Lepidoptera) (Sfbgly50). The active site of this enzyme was modeled based on its sequence and on crystallographic data of similar enzymes. Energy of noncovalent interactions in transition state between Sfbgly50 amino acids and glycone hydroxyls was determined. Sfbgly50 residue E451 seems to be a key residue in determining b-glycosidase preference for glucosides vs. galactosides based on the following data: (a)The energy of the noncovalent interaction between glycone equatorial hydroxyl 4 with E451 in the transition state is about 60% higher than its interaction with Q39. (b) The energy of the E451-hydroxyl 4 interaction decreases more than the Q39-hydroxyl 4 interaction when hydroxyl 4 is changed from equatorial to axial position. (c) A Sfbgly50 mutant, where E451 was substituted by A, hydrolyzes galactosides faster than glucosides. It was also shown that glycone hydroxyl 6 interacts favorably with Q39, but not with E451, probably due to steric hindrance. These interactions result in the b-glycosidase hydrolyzing fucosides (6-deoxygalactosides) faster than glucosides and galactosides.Keywords: b-glucosidase; glycoside hydrolase; enzyme specificity.The b-glycosidases from glycoside hydrolases family 1 are enzymes that remove monosaccharides from the nonreducing end of di-and/or oligo-saccharides. The nonreducing monosaccharide residue binds at the glycone subsite (subsite )1), whereas the rest of the substrate molecule accommodates at the aglycone subsite, which actually may correspond to several subsites (+1, +2, +3, etc.). The family 1 comprises a total of 213 sequenced b-glycosidases, from which 8 had their tertiary structure determined [1]. According to the system of glycoside hydrolases classification [1][2][3], all b-glycosidases from family 1 present the same tertiary structure (the (b/a) 8 barrel), they are configuration-retaining glycosidases and their catalytic activity depends on two glutamic acid residues, one positioned after the b strand 4 (proton donor) and the other after the b strand 7 (nucleophile). The family 1 comprises eight different EC numbers (3.2.1.21, b-glucosidase; 3.2.1.23, b-galactosidases; 3.2.1.25, b-mannosidases; 3.2.1.85, 6-phospho-b-galactosidase; 3.2.1.86, 6-phospho-b-glucosidase; 3.2.1.62, 3.2.1.108, lactase-phlorizin hydrolase and 3.2.3.1, myrosinase) catalyzing the hydrolysis of substrates presenting a variety of glycones (monosaccharides like glucose, galactose, fucose, mannose, 6-phospho-glucose and 6-phospho-ga...

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“…Hydrolysis of MUbglc (4-methylumbelliferyl b-D-glucopyranoside) was followed by MU fluorescence [22]. Kinetic parameters (k cat and K m ) were determined by using nine different substrate concentrations (0.1-8 mM); enzyme concentrations were 0.13 lM for R97M, 0.09 lM for Y331F and 0.62 lM for E187D.…”

Section: Kinetic Analysismentioning

confidence: 99%

The role of residues R97 and Y331 in modulating the pH optimum of an insect β‐glycosidase of family 1

Marana

Mendonça

Andrade

et al. 2003

European Journal of Biochemistry

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The activity of the digestive b-glycosidase from Spodoptera frugiperda (Sfbgly50, pH optimum 6.2) depends on E399 (pK a ¼ 4.9; catalytic nucleophile) and E187 (pK a ¼ 7.5; catalytic proton donor). Homology modelling of the Sfbgly50 active site confirms that R97 and Y331 form hydrogen bonds with E399. Site-directed mutagenesis showed that the substitution of R97 by methionine or lysine increased the E399 pK a by 0.6 or 0.8 units, respectively, shifting the pH optima of these mutants to 6.5. The substitution of Y331 by phenylalanine increased the pK a of E399 and E187 by 0.7 and 1.6 units, respectively, and displaced the pH optimum to 7.0. From the observed DpK a it was calculated that R97 and Y331 contribute 3.4 and 4.0 kJAEmol )1 , respectively, to stabilization of the charged E399, thus enabling it to be the catalytic nucleophile. The substitution of E187 by D decreased the pK a of residue 187 by 0.5 units and shifted the pH optimum to 5.8, suggesting that an electrostatic repulsion between the deprotonated E399 and E187 may increase the pK a of E187, which then becomes the catalytic proton donor. In short the data showed that a network of noncovalent interactions among R97, Y331, E399 and E187 controls the Sfbgly50 pH optimum. As those residues are conserved among the family 1 b-glycosidases, it is proposed here that similar interactions modulate the pH optimum of all family 1 b-glycosidases.Keywords: b-glycosidase; pK a values; pH optimum; sitedirected mutagenesis; Spodoptera frugiperda.The b-glycosidases from glycoside hydrolase family 1 are enzymes that remove monosaccharides from the nonreducing end of di-and/or oligosaccharides. According to the CAZy website this family comprises 422 sequenced b-glycosidases, of which the tertiary structure of 12 has been determined. Together with families 2, 10,17,26, 30, 35, 39, 42, 51, 53, 59, 72, 79 and 86 family 1 forms clan A, a group of families that shares structural and catalytic similarities [1]. All b-glycosidases of family 1 present the same tertiary structure [the (b/a) 8 barrel], they are configuration-retaining glycosidases and their catalytic activity depends on two glutamic acid residues, one positioned after b strand 4 and the other after b strand 7 [1]. These glutamic acids are very close inside the active site (about 4.5 Å apart) [2], and during the reaction the first glutamic acid acts as proton donor, and the second acts as a nucleophile. The catalytic nucleophile pK a is around 5.0 and the catalytic proton donor pK a is around 7.0 [3][4][5][6][7].A plot of b-glycosidase activity vs. pH presents a bell shape, indicating that in the pH optimum the catalytic nucleophile is deprotonated and the catalytic proton donor is protonated. Hence the branch of the curve below the pH optimum is determined mainly by the ionization of the catalytic nucleophile, whereas the catalytic ionization of the proton donor determines the branch above the pH optimum.As the b-glycosidase activity depends on the finely tuned ionization of the catalytic nucleophile and proton ...

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ß-glucosidases in the rice weevil, Sitophilus oryzae:Purification, properties, and activity levels in wheat- and legume-feeding strains

Cited by 38 publications

References 18 publications

The interplay between toxin-releasing β-glucosidase and plant iridoid glycosides impairs larval development in a generalist caterpillar, Grammia incorrupta (Arctiidae)

The interplay between toxin-releasing β-glucosidase and plant iridoid glycosides impairs larval development in a generalist caterpillar, Grammia incorrupta (Arctiidae)

The role of amino‐acid residues Q39 and E451 in the determination of substrate specificity of the Spodoptera frugiperdaβ‐glycosidase

The role of residues R97 and Y331 in modulating the pH optimum of an insect β‐glycosidase of family 1

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