A novel beta-1,4-xylanase from Trichoderma reesei

Maria Vrsanska; Matti Siika-aho; Markku Saloheimo; Peter Biely; Maija Tenkanen

A novel beta-1,4-xylanase from Trichoderma reesei

Maria Vrsanska, Matti Siika-aho, Markku Saloheimo, Peter Biely, Maija Tenkanen

Research output: Contribution to conference › Conference article › Scientific

Abstract

A novel xylanase was purified from xylan-spent culture fluid of the fungus Trichoderma reesei RUT-30. The novel xylanase, named XYL IV, exhibited catalytic properties incompatible with previously described endo--xylanases of Trichoderma reesei, XYL I and XYL II [1,2], XYL III [3] and xylan-hydrolyzing EG I [4]. The enzyme was discovered on the basis of its ability to attack aldotetrahexenuronic acid (HexA-2Xyl-4Xyl-4Xyl) releasing xylopyranosyl residue from the reducing end. XYL IV showed lower affinity toward glucuronoxylan than to acetylated beechwood glucuronoxylan and rhodymenan. Valuble information on the substrate-binding site of XYL IV was obtained by following the bond cleavage frequencies and kinetic parameters k0 /Km of [1-3H]-reducing end labeled xylooligosaccharides. Xyl2 was hydrolyzed by three orders of magnitude slower than longer xylooligosaccharides. Xyl3 and Xyl4 were found to be better enzyme substrates than Xyl5. Regardless, the substrate concentration, all three [1-3H]-xylooligosaccharides were exclusively cleaved at the first glycosidic linkage from the reducing end, to liberate [1-3H]-xylose as the only radioactive product. All experimental evidence suggests that the substrate-binding site of the Xyl IV is composed of three subsites, -II, -I and +I, with a serious steric barrier at the position +II. Degradation of methyl xylotrioside by XYL IV showed, that the hydrolysis leads exclusively to the liberation of the -anomer of Xyl2, which is slowly mutarotated to the -anomer. The time course of signals intensities of the -anomer and -anomer of Xyl2 confirmed that XYL IV is a retaining glycosyl hydrolase sign a double displacement reaction mechanism of hydrolysis of glycosidic linkage. [1] M. Tenkanen et al. Enzyme Microb. Technol. 14 (1992) 566-574. [2] P. Biely et al. In: Trichoderma reesei cellulases and other hydrolases (P. Souminen and T. Reinikainen, eds.). Foundation for Biotechnical and Industrial Fermentation Research, Helsinki, Vol. 8, (1993) 125-135. [3] J.Xu et al. Appl. Microbiol. Biotechnol. 49 (1998) 718-724. [4] P. Biely et al. Eur. J. Biochem. 200 (1991) 157-163.

Original language	English
Number of pages	1
Publication status	Published - 2003
MoE publication type	Not Eligible
Event	Molecular, Biochemical, Structural and Genetic Aspects of Carbohydrate-Modifying Enzymes - Galway, Ireland Duration: 14 Apr 2003 → 15 Apr 2003

Conference

Conference	Molecular, Biochemical, Structural and Genetic Aspects of Carbohydrate-Modifying Enzymes
Country/Territory	Ireland
City	Galway
Period	14/04/03 → 15/04/03

Cite this

@conference{5bdc670a57f34e809fd0df75517b3b3d,

title = "A novel beta-1,4-xylanase from Trichoderma reesei",

abstract = "A novel xylanase was purified from xylan-spent culture fluid of the fungus Trichoderma reesei RUT-30. The novel xylanase, named XYL IV, exhibited catalytic properties incompatible with previously described endo--xylanases of Trichoderma reesei, XYL I and XYL II [1,2], XYL III [3] and xylan-hydrolyzing EG I [4]. The enzyme was discovered on the basis of its ability to attack aldotetrahexenuronic acid (HexA-2Xyl-4Xyl-4Xyl) releasing xylopyranosyl residue from the reducing end. XYL IV showed lower affinity toward glucuronoxylan than to acetylated beechwood glucuronoxylan and rhodymenan. Valuble information on the substrate-binding site of XYL IV was obtained by following the bond cleavage frequencies and kinetic parameters k0 /Km of [1-3H]-reducing end labeled xylooligosaccharides. Xyl2 was hydrolyzed by three orders of magnitude slower than longer xylooligosaccharides. Xyl3 and Xyl4 were found to be better enzyme substrates than Xyl5. Regardless, the substrate concentration, all three [1-3H]-xylooligosaccharides were exclusively cleaved at the first glycosidic linkage from the reducing end, to liberate [1-3H]-xylose as the only radioactive product. All experimental evidence suggests that the substrate-binding site of the Xyl IV is composed of three subsites, -II, -I and +I, with a serious steric barrier at the position +II. Degradation of methyl xylotrioside by XYL IV showed, that the hydrolysis leads exclusively to the liberation of the -anomer of Xyl2, which is slowly mutarotated to the -anomer. The time course of signals intensities of the -anomer and -anomer of Xyl2 confirmed that XYL IV is a retaining glycosyl hydrolase sign a double displacement reaction mechanism of hydrolysis of glycosidic linkage. [1] M. Tenkanen et al. Enzyme Microb. Technol. 14 (1992) 566-574. [2] P. Biely et al. In: Trichoderma reesei cellulases and other hydrolases (P. Souminen and T. Reinikainen, eds.). Foundation for Biotechnical and Industrial Fermentation Research, Helsinki, Vol. 8, (1993) 125-135. [3] J.Xu et al. Appl. Microbiol. Biotechnol. 49 (1998) 718-724. [4] P. Biely et al. Eur. J. Biochem. 200 (1991) 157-163.",

author = "Maria Vrsanska and Matti Siika-aho and Markku Saloheimo and Peter Biely and Maija Tenkanen",

year = "2003",

language = "English",

note = "Molecular, Biochemical, Structural and Genetic Aspects of Carbohydrate-Modifying Enzymes ; Conference date: 14-04-2003 Through 15-04-2003",

}

TY - CONF

T1 - A novel beta-1,4-xylanase from Trichoderma reesei

AU - Vrsanska, Maria

AU - Siika-aho, Matti

AU - Saloheimo, Markku

AU - Biely, Peter

AU - Tenkanen, Maija

PY - 2003

Y1 - 2003

N2 - A novel xylanase was purified from xylan-spent culture fluid of the fungus Trichoderma reesei RUT-30. The novel xylanase, named XYL IV, exhibited catalytic properties incompatible with previously described endo--xylanases of Trichoderma reesei, XYL I and XYL II [1,2], XYL III [3] and xylan-hydrolyzing EG I [4]. The enzyme was discovered on the basis of its ability to attack aldotetrahexenuronic acid (HexA-2Xyl-4Xyl-4Xyl) releasing xylopyranosyl residue from the reducing end. XYL IV showed lower affinity toward glucuronoxylan than to acetylated beechwood glucuronoxylan and rhodymenan. Valuble information on the substrate-binding site of XYL IV was obtained by following the bond cleavage frequencies and kinetic parameters k0 /Km of [1-3H]-reducing end labeled xylooligosaccharides. Xyl2 was hydrolyzed by three orders of magnitude slower than longer xylooligosaccharides. Xyl3 and Xyl4 were found to be better enzyme substrates than Xyl5. Regardless, the substrate concentration, all three [1-3H]-xylooligosaccharides were exclusively cleaved at the first glycosidic linkage from the reducing end, to liberate [1-3H]-xylose as the only radioactive product. All experimental evidence suggests that the substrate-binding site of the Xyl IV is composed of three subsites, -II, -I and +I, with a serious steric barrier at the position +II. Degradation of methyl xylotrioside by XYL IV showed, that the hydrolysis leads exclusively to the liberation of the -anomer of Xyl2, which is slowly mutarotated to the -anomer. The time course of signals intensities of the -anomer and -anomer of Xyl2 confirmed that XYL IV is a retaining glycosyl hydrolase sign a double displacement reaction mechanism of hydrolysis of glycosidic linkage. [1] M. Tenkanen et al. Enzyme Microb. Technol. 14 (1992) 566-574. [2] P. Biely et al. In: Trichoderma reesei cellulases and other hydrolases (P. Souminen and T. Reinikainen, eds.). Foundation for Biotechnical and Industrial Fermentation Research, Helsinki, Vol. 8, (1993) 125-135. [3] J.Xu et al. Appl. Microbiol. Biotechnol. 49 (1998) 718-724. [4] P. Biely et al. Eur. J. Biochem. 200 (1991) 157-163.

AB - A novel xylanase was purified from xylan-spent culture fluid of the fungus Trichoderma reesei RUT-30. The novel xylanase, named XYL IV, exhibited catalytic properties incompatible with previously described endo--xylanases of Trichoderma reesei, XYL I and XYL II [1,2], XYL III [3] and xylan-hydrolyzing EG I [4]. The enzyme was discovered on the basis of its ability to attack aldotetrahexenuronic acid (HexA-2Xyl-4Xyl-4Xyl) releasing xylopyranosyl residue from the reducing end. XYL IV showed lower affinity toward glucuronoxylan than to acetylated beechwood glucuronoxylan and rhodymenan. Valuble information on the substrate-binding site of XYL IV was obtained by following the bond cleavage frequencies and kinetic parameters k0 /Km of [1-3H]-reducing end labeled xylooligosaccharides. Xyl2 was hydrolyzed by three orders of magnitude slower than longer xylooligosaccharides. Xyl3 and Xyl4 were found to be better enzyme substrates than Xyl5. Regardless, the substrate concentration, all three [1-3H]-xylooligosaccharides were exclusively cleaved at the first glycosidic linkage from the reducing end, to liberate [1-3H]-xylose as the only radioactive product. All experimental evidence suggests that the substrate-binding site of the Xyl IV is composed of three subsites, -II, -I and +I, with a serious steric barrier at the position +II. Degradation of methyl xylotrioside by XYL IV showed, that the hydrolysis leads exclusively to the liberation of the -anomer of Xyl2, which is slowly mutarotated to the -anomer. The time course of signals intensities of the -anomer and -anomer of Xyl2 confirmed that XYL IV is a retaining glycosyl hydrolase sign a double displacement reaction mechanism of hydrolysis of glycosidic linkage. [1] M. Tenkanen et al. Enzyme Microb. Technol. 14 (1992) 566-574. [2] P. Biely et al. In: Trichoderma reesei cellulases and other hydrolases (P. Souminen and T. Reinikainen, eds.). Foundation for Biotechnical and Industrial Fermentation Research, Helsinki, Vol. 8, (1993) 125-135. [3] J.Xu et al. Appl. Microbiol. Biotechnol. 49 (1998) 718-724. [4] P. Biely et al. Eur. J. Biochem. 200 (1991) 157-163.

M3 - Conference article

T2 - Molecular, Biochemical, Structural and Genetic Aspects of Carbohydrate-Modifying Enzymes

Y2 - 14 April 2003 through 15 April 2003

ER -

A novel beta-1,4-xylanase from Trichoderma reesei

Abstract

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