Real-time Adsorption And Action Of Expansin Onto Cellulose And Lignin Films

Expansin and expansin-like proteins act synergistically with cellulases during lignocellulose hydrolysis.Therefore,it is very useful to use expansin for improving hydrolysis efficiency of lignocellulose and saving cost.In our research,we have expressed and purified Bacillus subtilis expansin (BsEXLX1) in Pichia pastoris system.Quartz crystal microbalance with dissipation (QCM-D) was used to investigate the real-time adsorption and desorption behavior of BsEXLX1 onto cellulose and lignin films under several conditions.Furthermore,a kinetic model was established to fit the adsorption process.The main results were shown as follows.(1) Preparation and function of recombinant BsEXLX1.BsEXLX1,which has 39.13% ?-sheet,was expressed through Pichia pastoris system and then purified by preparative chromatography.In this part,the Avicel was used as substrate to investigate the function of BsEXLX1.BsEXLX1 was found to be synergetic with EG for improving the efficiency of cellulose hydrolysis.Besides,we also investigated the structural changes of cellulose upon BsEXLX1 adsorption by combined methods including FTIR,NMR and XRD.BsEXLX1 adsorption decreased the crystallinity index,disrupted hydrogen bonding,and increased the surface area of cellulose,indicating greater accessibility of cellulase to the substrate.(2) The real-time adsorption behavior of BsEXLX1 onto cellulose films.Upon 30 min BsEXLX1 treatment,the roughness and hydrophilicity of cellulose film increased obviously.During the adsorption stage,the frequency decreased rapidly,while the dissipation increased due to the change of viscoelasticity.High temperatures increased the initial adsorption rate while reducing the maximum amount of BsEXLX1 adsorbed onto the cellulose.Non-ionic surfactants (polyethylene glycol 4000 and Tween 80) at low concentrations enhanced BsEXLX1 adsorption whereas high concentrations had the opposite effect.However,sodium dodecyl sulfate (SDS) inhibited adsorption at both low and high concentrations.At last,a kinetic model was established to fit the adsorption process.And the adsorption,desorption,and irreversible adsorption rate constants were calculated.(3) The real-time adsorption behavior of BsEXLX1 onto lignin films.By a combined method including Zeta potential,FTIR and gel permeation chromatography(GPC),the structures of KL,SLS and EHL were analyzed and compared.The QCM-D results showed that KL had the highest adsorption capacity among the three samples because of its high molar mass and hydrophobicity.Rising the temperature to 45?lead to the decrease in maximum adsorption and initial adsorption rate.And Tween 80 could inhibit the adsorption of BsEXLX1 even at 0.02 mM.Furthermore,Tween 80 could partly peel off lignin when the concentration was increased to 1 mM.Compared the model parameter with cellulose,we found lignin has stronger binding ability with BsEXLX1.