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PDBsum entry 3a0f
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References listed in PDB file
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Key reference
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Title
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The crystal structure of a xyloglucan-Specific endo-Beta-1,4-Glucanase from geotrichum sp. M128 xyloglucanase reveals a key amino acid residue for substrate specificity.
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Authors
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K.Yaoi,
H.Kondo,
A.Hiyoshi,
N.Noro,
H.Sugimoto,
S.Tsuda,
K.Miyazaki.
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Ref.
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Febs J, 2009,
276,
5094-5100.
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PubMed id
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Abstract
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Geotrichum sp. M128 possesses two xyloglucan-specific glycoside hydrolases
belonging to family 74, xyloglucan-specific endo-beta-1,4-glucanase (XEG) and
oligoxyloglucan reducing-end-specific cellobiohydrolase (OXG-RCBH). Despite
their similar amino acid sequences (48% identity), their modes of action and
substrate specificities are distinct. XEG catalyzes the hydrolysis of xyloglucan
polysaccharides in endo mode, while OXG-RCBH acts on xyloglucan oligosaccharides
at the reducing end in exo mode. Here, we determined the crystal structure of
XEG at 2.5 A resolution, and compared it to a previously determined structure of
OXG-RCBH. For the most part, the amino acid residues that interact with
substrate are conserved between the two enzymes. However, there are notable
differences at subsite positions -1 and +2. OXG-RCBH has a loop around the +2
site that blocks one end of the active site cleft, which accounts for its exo
mode of action. In contrast, XEG lacks a corresponding loop at this site,
thereby allowing binding to the middle of the main chain of the substrate. At
the -1 site in OXG-RCBH, Asn488 interacts with the xylose side chain of the
substrate, whereas the -1 site is occupied by Tyr457 in XEG. To confirm the
contribution of this residue to substrate specificity, Tyr457 was substituted by
Gly in XEG. The wild-type XEG cleaved the oligoxyloglucan at a specific site;
the Y457G variant cleaved the same substrate, but at various sites. Together,
the absence of a loop in the cleft and the presence of bulky Tyr457 determine
the substrate specificity of XEG.
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