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Primary cell walls and middle lamella of higher plants include homogalacturonan, the substrate for polygalacturonases. Seeking to enhance lignocellulose saccharification, chimeras between the endopolygalacturonase I from Chondrostereum (Stereum) purpureum(EndoPG-I) and family 3, 44, and 77 carbohydrate-binding modules (CBMs) from Hungateiclostridium thermocellum(Ht) or Ruminococcus flavefaciens(Rf) were constructed, expressed, and characterized. Chimeras presented similar KMvalues and pH/temperature optima as unfused EndoPG-I against citrus pectin, kcat/KMvalues 1.6, 1.7, and 1.3-fold higher for the HtCBM3-EndoPG, HtCBM44-EndoPG, and RtCBM77-EndoPG, respectively. Commercial enzyme cocktail supplementation with HtCBM44-EndoPG and RtCBM77-EndoPG increased reducing sugar release from untreated sugarcane bagasse by 35 and 25%, respectively. All chimeras increased reducing sugar release by 20–25% against orange bagasse compared with EndoPG-I or equimolar EndoPG-I/CBM mixtures. These results show that proximity between plant cell wall components in situ influences CBM-enzyme chimera activity and improves saccharification of lignocellulosic materials.

A new strain of Trichoderma reesei(teleomorph Hypocrea jecorina) with high cellulase production was obtained by exposing the spores from T. reeseiQM9414 to an ultraviolet light followed by selecting fast-growing colonies on plates containing CMC (1% w/v) as the carbon source. The mutant T. reeseiRP698 reduced cultivation period to 5 days and increased tolerance to the end-products of enzymatic cellulose digestion. Under submerged fermentation conditions, FPase, CMCase, and Avicelase production increased up to 2-fold as compared to the original QM9414 strain. The highest levels of cellulase activity were obtained at 27 °C after 72 h with Avicel®, cellobiose, and sugarcane bagasse as carbon sources. The temperature and pH activity optima of the FPase, CMCase, and Avicelase were approximately 60 °C and 5.0, respectively. The cellulase activity was unaffected by the addition of 140 mM glucose in the enzyme assay. When T. reeseiRP698 crude extract was supplemented by the addition of β-glucosidase from Scytalidium thermophilum, a 2.3-fold increase in glucose release was observed, confirming the low inhibition by the end-product of cellulose hydrolysis. These features indicate the utility of this mutant strain in the production of enzymatic cocktails for biomass degradation.