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Title
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X-ray crystallographic studies of family 11 xylanase Michaelis and product complexes: implications for the catalytic mechanism.
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Authors
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Q.Wan,
Q.Zhang,
S.Hamilton-Brehm,
K.Weiss,
M.Mustyakimov,
L.Coates,
P.Langan,
D.Graham,
A.Kovalevsky.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2014,
70,
11-23.
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PubMed id
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Abstract
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Xylanases catalyze the hydrolysis of plant hemicellulose xylan into
oligosaccharides by cleaving the main-chain glycosidic linkages connecting
xylose subunits. To study ligand binding and to understand how the pH constrains
the activity of the enzyme, variants of the Trichoderma reesei xylanase were
designed to either abolish its activity (E177Q) or to change its pH optimum
(N44H). An E177Q-xylohexaose complex structure was obtained at 1.15 Å
resolution which represents a pseudo-Michaelis complex and confirmed the
conformational movement of the thumb region owing to ligand binding.
Co-crystallization of N44H with xylohexaose resulted in a hydrolyzed xylotriose
bound in the active site. Co-crystallization of the wild-type enzyme with
xylopentaose trapped an aglycone xylotriose and a transglycosylated glycone
product. Replacing amino acids near Glu177 decreased the xylanase activity but
increased the relative activity at alkaline pH. The substrate distortion in the
E177Q-xylohexaose structure expands the possible conformational itinerary of
this xylose ring during the enzyme-catalyzed xylan-hydrolysis reaction.
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