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ABSTRACT: Glucuronoyl esterase is a novel carbohydrate esterase recently discovered in the cellulolytic system of the wood-rotting fungus
Schizophyllum commune on the basis of its ability to hydrolyze methyl ester of 4-O-methyl-d-glucuronic acid. This substrate was not fully corresponding to the anticipated function of the enzyme to hydrolyze esters
between xylan-bound 4-O-methyl-d-glucuronic acid and lignin alcohols occurring in plant cell walls. In this work we showed that the enzyme was capable of
hydrolyzing two synthetic compounds that mimic the ester linkages described in lignin-carbohydrate complexes, esters of 4-O-methyl-d-glucuronic and d-glucuronic acid with 3-(4-methoxyphenyl)propyl alcohol. A comparison of kinetics of hydrolysis of methyl and 3-(4-methoxyphenyl)propyl
esters indicated that the glucuronoyl esterase recognizes the uronic acid part of the substrates better than the alcohol type.
The catalytic efficiency of the enzyme was much higher with the ester of 4-O-methyl-d-glucuronic acid than with that of d-glucuronic acid. Examination of the action of glucuronoyl esterase on a series of methyl esters of 4-O-methyl-d-glucopyranuronosyl residues α-1,2-linked to xylose and several xylooligosaccharides suggested that the rate of deesterification
is independent of the character of the carbohydrate part glycosylated by the 4-O-methyl-d-glucuronic acid.
Archives of Microbiology 04/2012; 188(2):185-189. · 1.43 Impact Factor
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ABSTRACT: The gene encoding Schizophyllum commune glucuronoyl esterase was identified in the scaffold 17 of the genome, containing two introns of 50 bp and 48 bp, with a transcript sequence of 1179 bp. The gene was synthesized and cloned into Pichia pastoris expression vector pGAPZα to achieve constitutive expression and secretion of the recombinant enzyme in soluble active form. The purified protein was 53 kD with glycosylation and had an acidic pI of 3.7. Activity analysis on several uronic acids and their derivatives suggests that the enzyme recognized only esters of 4-O-methyl-D-glucuronic acid derivatives, even with a 4-nitrophenyl aglycon but did not hydrolyze the ester of D-galacturonic acid. The kinetic values were K(m) 0.25 mM, V(max) 16.3 μM·min(-1), and k(cat) 9.27 s(-1) with 4-nitrophenyl 2-O-(methyl 4-O-methyl-α-D-glucopyranosyluronate)-β-D-xylopyranoside as the substrate.
Biotechnology research international. 01/2012; 2012:951267.
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ABSTRACT: Three acetyl esterases (AcEs) from the saprophytic bacteria Cellvibrio japonicus and Clostridium thermocellum, members of the carbohydrate esterase (CE) family 2, were tested for their activity against a series of model substrates including partially acetylated gluco-, manno- and xylopyranosides. All three enzymes showed a strong preference for deacetylation of the 6-position in aldohexoses. This regioselectivity is different from that of typical acetylxylan esterases (AcXEs). In aqueous medium saturated with vinyl acetate, the CE-2 enzymes catalyzed transacetylation to the same position, i.e., to the primary hydroxyl group of mono- and disaccharides. Xylose and xylooligosaccharides did not serve as acetyl group acceptors, therefore the CE-2 enzymes appear to be 6-O-deacetylases.
FEBS letters 12/2009; 584(3):543-8. · 3.54 Impact Factor
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ABSTRACT: Glucuronoyl esterases are enzymes involved in microbial plant cell-wall degradation. In this study we purified and characterized two recombinant Phanerochaete chrysosporium glucuronoyl esterases, PcGE1 and PcGE2. The catalytic activity of these and previously described glucuronoyl esterases was investigated on new synthetic substrates, methyl esters of uronic acids and their glycosides, prepared by esterification with ethereal diazomethane.The data obtained indicate that the enzymes hydrolyzed efficiently not only esters of 4-O-methyl-D-glucuronic acid, but also methyl esters of D-glucuronic acid carrying a 4-nitrophenyl aglycon. Moreover, the fact that they did not recognize the 4-epimers of these compounds, the D-galacturonic acid derivatives, supports the hypothesis that these carbohydrate esterases attack ester linkages between 4-O-methyl-D-glucuronic acid of glucuronoxylan and lignin alcohols.
Bioscience Biotechnology and Biochemistry 11/2009; 73(11):2483-7. · 1.28 Impact Factor
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ABSTRACT: Glucuronoyl esterase is a novel carbohydrate esterase recently discovered in the cellulolytic system of the wood-rotting fungus Schizophyllum commune on the basis of its ability to hydrolyze methyl ester of 4-O-methyl-D-glucuronic acid. This substrate was not fully corresponding to the anticipated function of the enzyme to hydrolyze esters between xylan-bound 4-O-methyl-D-glucuronic acid and lignin alcohols occurring in plant cell walls. In this work we showed that the enzyme was capable of hydrolyzing two synthetic compounds that mimic the ester linkages described in lignin-carbohydrate complexes, esters of 4-O-methyl-D-glucuronic and D-glucuronic acid with 3-(4-methoxyphenyl)propyl alcohol. A comparison of kinetics of hydrolysis of methyl and 3-(4-methoxyphenyl)propyl esters indicated that the glucuronoyl esterase recognizes the uronic acid part of the substrates better than the alcohol type. The catalytic efficiency of the enzyme was much higher with the ester of 4-O-methyl-D-glucuronic acid than with that of D-glucuronic acid. Examination of the action of glucuronoyl esterase on a series of methyl esters of 4-O-methyl-D-glucopyranuronosyl residues alpha-1,2-linked to xylose and several xylooligosaccharides suggested that the rate of deesterification is independent of the character of the carbohydrate part glycosylated by the 4-O-methyl-D-glucuronic acid.
Archives of Microbiology 09/2007; 188(2):185-9. · 1.43 Impact Factor
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ABSTRACT: A method for the preparation of methyl 4-O-methyl-alpha-D-glucopyranuronate and its single crystal X-ray structure determination are reported. The molecule adopts an almost ideal (4)C1 ((degree)C3) conformation.
Molecules 02/2005; 10(1):251-8. · 2.39 Impact Factor
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ABSTRACT: Fragmentation mechanisms of electrospray ionization (ESI) mass spectrometry of aldobiuronic and pseudoaldobiuronic acid derivatives were elucidated by multistage mass spectrometric (MS(n), n = 2-5) measurements of selected ions. Characteristic under the conditions of ESI-MS analysis is the production of alkali metal (Na and K) cationized adducts. The probability the of locations of Na cations in per-O-methylated compounds was proved by quantum chemical calculations, using the Jaguar program. The most probably position of alkali metal attachment is the carboxy group of the methoxycarbonyl C-5 group of the uronic acid unit. Characteristic cleavages vary according the kind of O-derivatization. In most cases they take place on the acidic part of the dimer and at the interglycosidic oxygen atom. As a result, the criteria for the differentiation of aldobiouronic and pseudoaldobiouronic acids derivatives were elucidated.
Journal of Mass Spectrometry 01/2005; 39(12):1554-61. · 3.27 Impact Factor
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ABSTRACT: 4-Nitrophenyl 2-(4-O-methyl-α--glucopyranuronosyl)-β--xylopyranoside obtained on deesterification of 4-nitrophenyl 2-O-(methyl 4-O-methyl-α--glucopyranosyluronate)-β--xylopyranoside (Hirsch et al., Carbohydr. Res. 310, 145–149, 1998) was found to be an excellent substrate for the measurement of hemicellulolytic α-glucuronidase activity. A new precise α-glucuronidase assay was developed by coupling the α-glucuronidase-catalyzed formation of 4-nitrophenyl β--xylopyranoside with its efficient hydrolysis by β-xylosidase. A recombinant strain of Saccharomyces cerevisiae, harboring and expressing the β-xylosidase gene xlnD of Aspergillus niger under control of the alcohol dehydrogenase II promoter on a multicopy plasmid, was used as a source of β-xylosidase. The activity values of β-xylosidase in the assay required to achieve a steady-state rate of 4-nitrophenol formation shortly after starting the α-glucuronidase reaction were obtained both experimentally and by calculation using the kinetics of coupled enzyme reactions.
Analytical Biochemistry.
