Xylanase-aided Enzymatic Treatment To Facilitate Lignocellulose Deconstruction,Modification And Xylan Fluorescent Blotting

With the advances in xylanase-aided treatment on lignocellulose bioconversion,we introduced several xylanase-mediated enzymatic approaches to facilitate cellulose nanofibrillation,cellulos e-based composite modification and lignocellulose saccharification at high solids loading accordingly.In addition,a thermostable xylan specific binding domain from thermostable Thermotoga thermarum DSM 5069 xylanase?Xyn10A?was cloned and characterized as a potential molecular probe to monitor the interfacial xylan present on cellulosic fibers.This project aimed at realizing the conversion of lignocellulose to high value-added products?energy and materials?more efficiently and accurately.?1?Although it has been shown that cellulosic fiber with endoglucanase pretreatment?EG,which randomly cleavages the ce llulose?-1,4 linkages at less organized region?could improve the downstream cellulose fibrillation,the extent of such improvement was still quite limited.In the work reported here,we assessed the potential benefits of supplementation of xylanases to endoglucanase to facilitate the biological pretreatment of bleached Kraft pulp from hardwood and softwood,respectively,for enhancing the downstream cellulose nanofibrillation.Results showed that the synergistic cooperation between endoglucanase and certain xylanase?Biobrite?could efficiently“open up”the hardwood Kraft pulp with limited carbohydrates degradation?<7%?,stating that the binary combination of EG and Biobrite was amendable to facilate cellulose nanofibrillation.This study also provided a solid founda tion for our following research supported by the advantages of xylanase-assisted enzymatic treatment.?2?Xylanases are able to specifically hydrolyze xylan component coated on cellulose fiber without compromising the structural advantages of cellulose.The xylanase treatment strategy was therefore conducted to assess the structural role of xylan that played in the regenerated cellulose/xylan composite films,and exploring its technical feasibility for the selective surface modifications at the same time.Results showed that the distribution of xylan component on the surface was highly heterogenous.The coated xylan component was highly accessible to xylanase enzyme,e.g.26.6%and 32.3%xylan was released with the addition of 2 mg/g and 5 mg/g xylanase,re spectively.After xylanase treatment,the surface of the composite films was highly etched/roughed at nanoscale.The extent of surface etching/roughing of the composite films could be facilely controlled by manipulating xylanase loading.?3?Beyond the challenges of researching viable alternative cellulosic biofuels is that of bringing the biofuel production from lab scale to commercial scale.Saccharification of cellulosic component at high solids loading is therefore required to meet the production demand.To facilitate the extend of hydrolysis yield,a temperature-elevated two-stage hydrolysis,including xylan“coat”removal at high-temperature by thermostable xylanase?Xyn10A?from Thermotoga thermarum DSM 5069 followed with saccharification step by commercial cellulase,was introduced to improve biomass deconstruction in the4^th chapter.Results showed that high-temperature xylanase treatment considerably increased cellulose accessibility/hydrolyzability towards cellulases,with smoothed fiber surface mo rphology.It appeared that the increased temperature during thermostable xylanase treatment facilitated biomass slurry viscosity reduction,which exhibited more benefits during hydrolysis of various steam pretreated substrates at increased solid content?up to 10%w/w?.?4?Specific techniques to localize and visualize the spatial distribution of the major polymer compositions of lignocellulose is crucial to better understand and control the fine processing of lignocellulosic biomass.In the last chapter,a xylan specific binding domain from thermostable Thermotoga thermarum DSM5069 xylanase?Xyn10A?N-terminal domain?N1-N2?was cloned and characterized as a potential molecular probe to monitor the interfacial xylan of cellulosic fibers.The results sho wed that N1-N2 could selectively interact with both insoluble and soluble xylanolytic substrates?no affinity with either crystalline or amorphous cellulose?.When the tagged thermostable xylan binding module N1-N2 was incubated with commercial Kraft pulps derived from hardwood?17.2%xylan?and softwood?8.4%xylan?,respectively,the distinct binding patterns were observed under confocal microscopy.We believe that this tracking approach could decrease costs associated with lignocellulosic biomass processes and expand our understanding of biofuels and papermaking productions.