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Glycoside hydrolases of this family hydrolyze β-1,4-linkages of various glucans. With the exception of Cel74 from Thermotoga maritima, all biochemically characterized enzymes are specific toward xyloglucans and/or xyloglucan-oligosaccharides (reviewed in [1]). Cel74 from Thermotoga maritima exhibits the highest activity on barley β-glucan, with relative activity of 20% toward xyloglucan [2]. A wide diversity in the modes of action by GH74 enzymes has been reported. "Oligoxyloglucan reducing end-specific cellobiohydrolase (OXG-RCBH, EC 3.2.1.150)" from Geotrichum sp. M128 [3] and "oligoxyloglucan reducing end-specific xyloglucanobiohydrolase (OREX)" from Emericella nidulans (formerly known as Aspergillus nidulans) [4] are active on only xyloglucan oligosaccharides and have essentially no ability to degrade xyloglucan polysaccharides. They release oligosaccharides with two glucose units from non-reducing end of xyloglucan oligosaccharides. On the other hand, GH74 enzymes designated as xyloglucanase; xyloglucan specific endo-β-1,4-glucanases: XEG; and xyloglucan hydrolases: Xgh, (EC 3.2.1.151), exhibit endo-type activity on xyloglucan from tamarind seed, a readily available and well-investigated xyloglucan [5]. Many GH74 xyloglucanases hydrolyze the glycosidic linkage of unbranched glucose residues, but several members including Geotrichum sp. OXG-RCBH [3], E. nidulans OREX [4], and Hypocrea jecorina (formerly known as Trichoderma reesei) Cel74A [6] accommodate side-chain xylose residues at subsite -1 of the active site. As first elucidated by Matsuzawa, Saito, and Yaoi [7], some GH74 members exhibit processivity (multiple catalytic events before chain dissociation) in xyloglucan hydrolysis, which is related to active-site length and the presence of key aromatic residues, and which follows protein phylogeny [1].
Crystal structure of OXG-RCBH demonstrated that Asp35 and Asp465 are located in the middle of the binding cleft, and their crucial roles in hydrolytic activity were experimentally confirmed by site-directed mutagenesis [9]. However, their identities as general acid and general base were not assigned. The corresponding Asp residues in Xgh74A from Clostridium thermocellum are nicely located between subsites -1 and +1 in the complex structure with xyloglucan-derived oligosaccharides [10].
Three-dimensional structures
Overall structures of GH74 enzymes consist of a tandem repeat of two seven-bladed β-propeller domains. The two domains form a substrate binding cleft at the interface. The catalytic residues are located in the middle of this cleft. One side of the binding cleft of OXG-RCBH is blocked by a so-called 'exo-loop' which is found only in exo-acting enzymes in this family [9]. A crystal structure of a complex with xyloglucan-derived oligosaccharides elucidated the interaction with the side-chains of the substrate by these enzymes [10]. A number of GH74 complexes with large xyloglucan oligosaccharides is now known [1].
Takada G, Kawagushi T, Yoneda T, Kawasaki M, Sumitani JI, and Arai M. Molecular cloning and expression of the celluloytic system of Aspergillus aculeatus, p. 364-373. In Ohmiya K, Hayashi K, Sakka K, Kobayashi Y, Karita S, and Kimura T (ed.), Genetics, biochemistry and ecology of cellulose degradation. 1999 Uni Publishers, Tokyo, Japan, ISBN 4-946450-17-3. [Takada1999]
