Crystallographic evidence for substrate ring distortion and protein conformational changes during catalysis in cellobiohydrolase Cel6A from Trichoderma reesei

Jin-yu Zou, Gerald Kleywegt, Jerry Ståhlberg, Hugues Driguez, Wim Nerinckx, Marc Claeyssens, Anu Koivula, Tuula Teeri, Alwyn Jones (Corresponding Author)

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144 Citations (Scopus)


Background: Cel6A is one of the two cellobiohydrolases produced by Trichoderma reesei. The catalytic core has a structure that is a variation of the classic TIM barrel. The active site is located inside a tunnel, the roof of which is formed mainly by a pair of loops.

Results: We describe three new ligand complexes. One is the structure of the wild-type enzyme in complex with a nonhydrolysable cello-oligosaccharide, methyl 4-S-β-cellobiosyl-4-thio-β-cellobioside (Glc)2-S-(Glc)2, which differs from a cellotetraose in the nature of the central glycosidic linkage where a sulphur atom replaces an oxygen atom. The second structure is a mutant, Y169F, in complex with the same ligand, and the third is the wild-type enzyme in complex with m-iodobenzyl β-D-glucopyranosyl-β(1,4)-D-xylopyranoside (IBXG).

Conclusions: The (Glc)2-S-(Glc)2 ligand binds in the -2 to +2 sites in both the wild-type and mutant enzymes. The glucosyl unit in the -1 site is distorted from the usual chair conformation in both structures. The IBXG ligand binds in the -2 to +1 sites, with the xylosyl unit in the -1 site where it adopts the energetically favourable chair conformation. The -1 site glucosyl of the (Glc)2-S-(Glc)2 ligand is unable to take on this conformation because of steric clashes with the protein. The crystallographic results show that one of the tunnel-forming loops in Cel6A is sensitive to modifications at the active site, and is able to take on a number of different conformations. One of the conformational changes disrupts a set of interactions at the active site that we propose is an integral part of the reaction mechanism.
Original languageEnglish
Pages (from-to)1035-1045
Number of pages11
Issue number9
Publication statusPublished - 1999
MoE publication typeA1 Journal article-refereed


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