The crystal structure of a xyloglucan-specific endo-b-1,4-glucanase from Geotrichum sp. M128 xyloglucanase reveals a key amino acid residue for substrate specificity
Geotrichum sp. M128 possesses two xyloglucan-specific glycoside hydrolases belonging to family 74, xyloglucan-specific endo-b-1,4-glucanase (XEG) and oligoxyloglucan reducing-end-specific cellobiohydrolase (OXG-RCBH). Despite their similar amino acid sequences (48% identity), their modes of action a...
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Published in | The FEBS journal Vol. 276; no. 18; pp. 5094 - 5100 |
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Main Authors | , , , , , , |
Format | Journal Article |
Language | English |
Published |
01.09.2009
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Subjects | |
Online Access | Get full text |
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Summary: | Geotrichum sp. M128 possesses two xyloglucan-specific glycoside hydrolases belonging to family 74, xyloglucan-specific endo-b-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 Aa 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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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 content type line 23 ObjectType-Feature-2 |
ISSN: | 1742-464X |
DOI: | 10.1111/j.1742-4658.2009.07205.x |