Diverse substrate recognition mechanism revealed by Thermotoga maritima Cel5A structures in complex with cellotetraose, cellobiose and mannotriose

Tracing Determinants of Dual Substrate Specificity in Glycoside Hydrolase Family 5

“…The strong effect of the His-95 mutation in reducing glucanase activity can be explained by its interaction with the Ϫ1 subsite OH-C2 when the OH is in the equatorial configuration in the glucan substrate, whereas this interaction does not appear to occur for the axial conformation found in the mannan substrate. A recently released structure of Cel5A_Tma in complex with different sugar moieties confirms our model and supports our interpretations (34). Although there are no reports in the literature for enzymatic activity enhancement for mannan hydrolysis, the largest improvements for glucan and xylan hydrolysis to date are ϳ80% (35) and ϳ300% (36), respectively.…”

Biochemical Characterization and Structural Analysis of a Bifunctional Cellulase/Xylanase from Clostridium thermocellum

“…Site-directed mutagenesis studies (Table 2) indicate that Glu-209 and Glu-314 are these two catalytic residues. In addition, Arg-124, Asn-208, His-268, and Trp-347 are strictly conserved and posited similarly in the active site compared with those in other GH5 enzymes (10,11,25) (Fig. 3C).…”

“…In addition, one ␣ helix at the N terminus is located at the bottom of the barrel, and one 3 10 helix is located at the top of barrel.…”

“…In addition, we determined the crystal structures of apo-and ligand-bound CtCel5E and subsequently carried out site-directed mutagene-sis to confirm the roles of the active-site amino acid residues. Structural comparison between CtCel5E and its homologue, TmCel5A (9,10), which is a bifunctional Cel5A from Thermotoga maritima and can degrade cellulose and mannan (11), provides insights into substrate recognition and may lead to CtCel5E functional improvement by protein engineering. Thus, our study is of general interest because engineered multiple-function enzymes can be a convenient, economical, and efficient tool in biofuel production because of their broad substrate specificities for processing plant feedstock (12)(13)(14)(15).…”

Lignocellulases: a review of emerging and developing enzymes, systems, and practices

“…On account of the obvious benefits reported in the literature, scientists have consistently investigated the gene (Yagüe et al 1990;Zverlov et al 2003;Koeck et al 2013), fusion/modification of enzymes (Ciolacu et al 2010;Lee et al 2010;Ye et al 2010;Lee et al 2011;Nakashima et al 2014), and the optimal growth (Islam et al 2013;Reed et al 2014) of these useful microbes to harness their inherent benefits. For example, the biochemical and biophysical characteristics of multimodular enzymes from Clostridium thermocellum (Zverlov et al 2005;Tachaapaikoon et al 2012;Brunecky et al 2012;Hirano et al 2013;Yuan et al 2015) and Thermotoga maritima (Chhabra et al 2002;Carvalho et al 2004;Pereira et al 2010;Wu et al 2011) have been reported.…”