Date of Award
3-17-2020
Document Type
Dissertation
Degree Name
Molecular Biosciences, Ph.D.
First Advisor
Elizabeth Hood
Committee Members
Brett Savary; Gregory Phillips; Jianfeng Xu; John Howard
Call Number
LD 251 .A566d 2019 F35
Abstract
Enzymatic saccharification of ligno-cellulosic material is a key process in advanced biofuel production. Different enzymes are required in the enzymatic saccharification. To meet the large demand of these enzymes for industrial purposes, a transgenic maize seed expression system had been utilized in our laboratory to acquire adequate and cost-effective pure enzymes including β-1,4-endoglucanase, cellobiohydrolase I, cellobiohydrolase II and expansin. In this dissertation, the possibility of utilizing maize-produced recombinant cellulases in promoting the saccharification of ligno-cellulosic material was investigated. From the purification and basic characterization of these target enzymes to the identification of different inhibitors, this project has partially addressed the efficiency concerns of maize-produced recombinant cellulases and highlights advantages of the maize expression system. In general, this dissertation includes four parts: the identification of pinewood-derived cellulase inhibitors, the identification of maize native cellulase inhibitors, the characterization of recombinant expansin and the discussion of the three major activity assays involved throughout the entire work. In the first part, pinewood-derived inhibitors as a potential obstacle in using pinetree logging residues as the ligno-cellulosic feedstock were investigated. The result showed that oligomers composed of xylan, galactan, and mannan were the main inhibitors in the pinewood pre-hydrolysate. In the second part, a maize arabinoxylan was determined as the cellulase inhibitor, which was irremovable in the maize seed expression system due to its interaction with the target recombinant cellulases and its resistance to related hydrolysis enzymes. In the third part, we failed in the isolation and characterization of the target recombinant expansin due to the lack of specific identification tools. However, instead of expansin, a novel maize native β-glucosidase was discovered from maize seeds. In the last part, all enzymatic assays involved in this dissertation were compared and discussed based on the data obtained throughout the entire body of work. In conclusion, we confirmed that the maize seed expression system is able to produce pure recombinant cellulases for ligno-cellulosic material saccharification and allowed the identification of novel inhibitors of cellulase enzymes. Moreover, a novel β-glucosidase from seed was discovered and characterized.
Rights Management
This work is licensed under a Creative Commons Attribution 4.0 International License.
Recommended Citation
Fang, Hong, "Cellulolytic Enzymes from The Maize Seed Production System—Characterization, Inhibition, And Novel Activities" (2020). Student Theses and Dissertations. 415.
https://arch.astate.edu/all-etd/415
