Atomic Force Microscopy Study of Endoglucanases and Cellobiohydrolases on Native Cellulose Films

dc.contributor.advisorLipkowski, Jacek
dc.contributor.advisorRoscoe, Sharon
dc.contributor.authorQuirk, Amanda
dc.date.accessioned2012-03-20T19:40:58Z
dc.date.available2012-03-20T19:40:58Z
dc.date.copyright2012-03
dc.date.created2012-03-01
dc.date.issued2012-03-20
dc.degree.departmentDepartment of Chemistryen_US
dc.degree.grantorUniversity of Guelphen_US
dc.degree.nameDoctor of Philosophyen_US
dc.degree.programmeChemistryen_US
dc.description.abstractAtomic force microscopy was used to image the action of cellulolytic enzymes in situ on never-dried native cellulose films. Cellomonas fimi, CenA was used as a model enzyme for proof of concept experiments and for the identification of different enzyme action on different cellulose structures. Inactive and active Trichoderma reesei enzymes EGI and CBHI were studied to disentangle the action of the cellulose binding domain from the catalytic domain. A novel procedure, volume analysis, was developed to quantify changes in cellulose fibers as a result of this action. Volume analysis was used to compare fibers in different experiments (with different structural features and enzymes) regardless of where the change in the fiber occurred. The site-specific nature of cellulose-enzyme interactions is accessible using this analysis technique. Additionally, the reported volume change reflects a change in mass that is of interest for industrial purposes. From inactive CBHI action there was no distinguishable change between enzyme action on defect or crystalline regions of the cellulose fiber. From the active enzyme results a quantifiable degradation event was measured. Digestion was initially quick then after one hour the volume plateaued. The crystalline cellulose region plateaued at -20 ± 1% and the defect region at -31 ± 2%. The inactive EGI enzyme was found to have significant non-hydrolytic action on insoluble cellulose fibers. There was more significant swelling effect on the defect than the crystalline regions of the cellulose fiber. From the active EGI results a quantifiable degradation event was measured followed by swelling events. Degradation was initially quick with the total mass loss occurring within the first hour of the experiment. The volume then increased as the enzyme induced swelling of the fiber structure. The extent of degradation and swelling is structure limited with more disordered regions showing larger decreases in volume and predominantly crystalline regions showing mainly swelling events.en_US
dc.identifier.urihttp://hdl.handle.net/10214/3468
dc.language.isoenen_US
dc.publisherUniversity of Guelphen_US
dc.rights.licenseAll items in the Atrium are protected by copyright with all rights reserved unless otherwise indicated.
dc.subjectatomic force microscopyen_US
dc.subjectcelluloseen_US
dc.subjectendoglucanaseen_US
dc.subjectcellobiohydrolaseen_US
dc.subjectbiofuelen_US
dc.titleAtomic Force Microscopy Study of Endoglucanases and Cellobiohydrolases on Native Cellulose Filmsen_US
dc.typeThesisen_US

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