The mechanisms by which family 10 glycoside hydrolases bind decorated substrates

Pell , Gavin and Taylor, Edward J. and Gloster, Tracey M. and Turkenburg, Johan P. and Fontes, Carlos M. G. A. and Ferreira, Luis M.A. and Nagy, Tibor and Clark, Samantha J. and Davies, Gideon J. and Gilbert, Harry J. (2004) The mechanisms by which family 10 glycoside hydrolases bind decorated substrates. Journal of Biological Chemistry , 279 (10). pp. 9597-9605. ISSN 0021-9258

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Official URL: http://dx.doi.org/10.1074/jbc.M312278200

Abstract

Endo-{beta}-1,4-xylanases (xylanases), which cleave {beta}-1,4 glycosidic bonds in the xylan backbone, are important components of the repertoire of enzymes that catalyze plant cell wall degradation. The mechanism by which these enzymes are able to hydrolyze a range of decorated xylans remains unclear. Here we reveal the three-dimensional structure, determined by x-ray crystallography, and the catalytic properties of the Cellvibrio mixtus enzyme Xyn10B (CmXyn10B), the most active GH10 xylanase described to date. The crystal structure of the enzyme in complex with xylopentaose reveals that at the +1 subsite the xylose moiety is sandwiched between hydrophobic residues, which is likely to mediate tighter binding than in other GH10 xylanases. The crystal structure of the xylanase in complex with a range of decorated xylooligosaccharides reveals how this enzyme is able to hydrolyze substituted xylan. Solvent exposure of the O-2 groups of xylose at the +4, +3, +1, and -3 subsites may allow accommodation of the {alpha}-1,2-linked 4-O-methyl-D-glucuronic acid side chain in glucuronoxylan at these locations. Furthermore, the uronic acid makes hydrogen bonds and hydrophobic interactions with the enzyme at the +1 subsite, indicating that the sugar decorations in glucuronoxylan are targeted to this proximal aglycone binding site. Accommodation of 3'-linked L-arabinofuranoside decorations is observed in the -2 subsite and could, most likely, be tolerated when bound to xylosides in -3 and +4. A notable feature of the binding mode of decorated substrates is the way in which the subsite specificities are tailored both to prevent the formation of "dead-end" reaction products and to facilitate synergy with the xylan degradation-accessory enzymes such as {alpha}-glucuronidase. The data described in this report and in the accompanying paper (Fujimoto, Z., Kaneko, S., Kuno, A., Kobayashi, H., Kusakabe, I., and Mizuno, H. (2004) J. Biol. Chem. 279, 9606-9614) indicate that the complementarity in the binding of decorated substrates between the glycone and aglycone regions appears to be a conserved feature of GH10 xylanases. Pell, G., Taylor, E. J., Gloster, T. M., Turkenburg, J. P., Fontes, C., Ferreira, L. M. A., Nagy, T., Clark, S. J., Davies, G. J. & Gilbert, H. J.

Item Type:Article
Additional Information:Endo-{beta}-1,4-xylanases (xylanases), which cleave {beta}-1,4 glycosidic bonds in the xylan backbone, are important components of the repertoire of enzymes that catalyze plant cell wall degradation. The mechanism by which these enzymes are able to hydrolyze a range of decorated xylans remains unclear. Here we reveal the three-dimensional structure, determined by x-ray crystallography, and the catalytic properties of the Cellvibrio mixtus enzyme Xyn10B (CmXyn10B), the most active GH10 xylanase described to date. The crystal structure of the enzyme in complex with xylopentaose reveals that at the +1 subsite the xylose moiety is sandwiched between hydrophobic residues, which is likely to mediate tighter binding than in other GH10 xylanases. The crystal structure of the xylanase in complex with a range of decorated xylooligosaccharides reveals how this enzyme is able to hydrolyze substituted xylan. Solvent exposure of the O-2 groups of xylose at the +4, +3, +1, and -3 subsites may allow accommodation of the {alpha}-1,2-linked 4-O-methyl-D-glucuronic acid side chain in glucuronoxylan at these locations. Furthermore, the uronic acid makes hydrogen bonds and hydrophobic interactions with the enzyme at the +1 subsite, indicating that the sugar decorations in glucuronoxylan are targeted to this proximal aglycone binding site. Accommodation of 3'-linked L-arabinofuranoside decorations is observed in the -2 subsite and could, most likely, be tolerated when bound to xylosides in -3 and +4. A notable feature of the binding mode of decorated substrates is the way in which the subsite specificities are tailored both to prevent the formation of "dead-end" reaction products and to facilitate synergy with the xylan degradation-accessory enzymes such as {alpha}-glucuronidase. The data described in this report and in the accompanying paper (Fujimoto, Z., Kaneko, S., Kuno, A., Kobayashi, H., Kusakabe, I., and Mizuno, H. (2004) J. Biol. Chem. 279, 9606-9614) indicate that the complementarity in the binding of decorated substrates between the glycone and aglycone regions appears to be a conserved feature of GH10 xylanases. Pell, G., Taylor, E. J., Gloster, T. M., Turkenburg, J. P., Fontes, C., Ferreira, L. M. A., Nagy, T., Clark, S. J., Davies, G. J. & Gilbert, H. J.
Keywords:xylanases, gh10, plant cell wall degradation
Subjects:C Biological Sciences > C700 Molecular Biology, Biophysics and Biochemistry
Divisions:College of Science > School of Life Sciences
ID Code:6177
Deposited By: Edward Taylor
Deposited On:18 Sep 2012 12:31
Last Modified:13 Mar 2013 09:13

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