Abstract
Xylanase B, a member of subfamily 7 of the GH30 (glycoside hydrolase family 30) from Talaromyces cellulolyticus (TcXyn30B), is a bifunctional enzyme with glucuronoxylanase and xylobiohydrolase activities. In the present study, crystal structures of the native enzyme and the enzyme–product complex of TcXyn30B expressed in Pichia pastoris were determined at resolutions of 1.60 and 1.65 Å, respectively. The enzyme complexed with 22‐(4‐O‐methyl‐α‐d‐glucuronyl)‐xylobiose (U4m2X) revealed that TcXyn30B strictly recognizes both the C‐6 carboxyl group and the 4‐O‐methyl group of the 4‐O‐methyl‐α‐d‐glucuronyl side chain by the conserved residues in GH30‐7 endoxylanases. The crystal structure and site‐directed mutagenesis indicated that Asn‐93 on the β2‐α2‐loop interacts with the non‐reducing end of the xylose residue at subsite‐2 and is likely to be involved in xylobiohydrolase activity. These findings provide structural insight into the mechanisms of substrate recognition of GH30‐7 glucuronoxylanase and xylobiohydrolase.
Highlights
IntroductionXylanase B, a member of subfamily 7 of the GH30 (glycoside hydrolase family 30) from Talaromyces cellulolyticus (TcXyn30B), is a bifunctional enzyme with glucuronoxylanase and xylobiohydrolase activities
Xylanase B, a member of subfamily 7 of the GH30 from Talaromyces cellulolyticus (TcXyn30B), is a bifunctional enzyme with glucuronoxylanase and xylobiohydrolase activities
The crystal structure and site-directed mutagenesis indicated that Asn-93 on the b2-a2-loop interacts with the non-reducing end of the xylose residue at subsite-2 and is likely to be involved in xylobiohydrolase activity
Summary
Xylanase B, a member of subfamily 7 of the GH30 (glycoside hydrolase family 30) from Talaromyces cellulolyticus (TcXyn30B), is a bifunctional enzyme with glucuronoxylanase and xylobiohydrolase activities. The crystal structure and site-directed mutagenesis indicated that Asn-93 on the b2-a2-loop interacts with the non-reducing end of the xylose residue at subsite-2 and is likely to be involved in xylobiohydrolase activity. These findings provide structural insight into the mechanisms of substrate recognition of GH30-7 glucuronoxylanase and xylobiohydrolase. Crystal structures of GH30-8, such as EcXynA from Dickeya chrysanthemi (formerly Erwinia chrysanthemi) and BsXynC from Bacillus subtilis, have revealed that the enzymes consist of a (b/ a)8-barrel with an obligatory side-associated, ninestranded, aligned b-sandwich [1,6].
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