Rice straw (RS) is an abundant, readily available agricultural waste, which shows promise as a potential feedstock for Asian ethanol production. To enhance release of glucose by enzymatic hydrolysis, RS was pretreated with aqueous ammonia (27% w/w) at two pretreatment temperatures: room temperature and 60°C. Statistical analysis indicated similarity of enzymatic glucose production at both pretreatment temperatures after 3-day incubation. Chemical composition, FTIR, and EDX analyses confirmed the retention of glucan and xylan in the pretreated solid, but significant reduction of lignin (60.7% removal) and silica. SEM analysis showed the disorganized surfaces and porosity of the pretreated RS fibers, thus improving cellulose accessibility for cellulase. The crystallinity index increased from 40.5 to 52.3%, indicating the higher exposure of cellulose. With 10% (w/v) solid loadings of pretreated RS, simultaneous saccharification and fermentation yielded a final ethanol concentration of 24.6 g/L, corresponding to 98% of maximum theoretical yield. Taken together, aqueous ammonia pretreatment is an effective method to generate highly digestible pretreated RS for bioethanol production and demonstrates potential application in biorefinery industry.

The axy43A gene encoding the intracellular trifunctional xylanolytic enzyme from Paenibacillus curdlanolyticus B-6 was cloned and expressed in Escherichia coli . Recombinant PcAxy43A consisting of a glycoside hydrolase family 43 and a family 6 carbohydrate-binding module exhibited endo-xylanase, β-xylosidase, and arabinoxylan arabinofuranohydrolase activities. PcAxy43A hydrolyzed xylohexaose and birch wood xylan to release a series of xylooligosaccharides, indicating that PcAxy43A contained endo-xylanase activity. PcAxy43A exhibited β-xylosidase activity toward a chromogenic substrate, p -nitrophenyl-β- d -xylopyranoside, and xylobiose, while it preferred to hydrolyze long-chain xylooligosaccharides rather than xylobiose. In addition, surprisingly, PcAxy43A showed arabinoxylan arabinofuranohydrolase activity; that is, it released arabinose from both singly and doubly arabinosylated xylose, α- l -Ara f -(1→2)- d -Xyl p or α- l -Ara f -(1→3)- d -Xyl p and α- l -Ara f -(1→2)-[α- l -Ara f -(1→3)]-β- d -Xyl p . Moreover, the combination of PcAxy43A and P. curdlanolyticus B-6 endo-xylanase Xyn10C greatly improved the efficiency of xylose and arabinose production from the highly substituted rye arabinoxylan, suggesting that these two enzymes function synergistically to depolymerize arabinoxylan. Therefore, PcAxy43A has the potential for the saccharification of arabinoxylan into simple sugars for many applications. IMPORTANCE In this study, the glycoside hydrolase 43 (GH43) intracellular multifunctional endo-xylanase, β-xylosidase, and arabinoxylan arabinofuranohydrolase (AXH) from P. curdlanolyticus B-6 were characterized. Interestingly, PcAxy43A AXH showed a new property that acted on both the C(O)-2 and C(O)-3 positions of xylose residues doubly substituted with arabinosyl, which usually obstruct the action of xylanolytic enzymes. Furthermore, the studies here show interesting properties for the processing of xylans from cereal grains, particularly rye arabinoxylan, and show a novel relationship between PcAxy43A and endo-xylanase Xyn10C from strain B-6, providing novel metabolic potential for processing arabinoxylans into xylose and arabinose.