Combinatorial biosynthetic pathway engineering for microbial production of biofuels

To compete in a market dominated by fossil fuels, biofuels must be economically competitive and also offer the variety of molecules and compounds which are currently derived from fossil fuels. This thesis offers potential strategies for biofuels to be both economically competitive and diverse. Effective and economical production of biofuels comes from the optimization of the biosynthetic pathway. We investigated and developed new combinatorial strategies for the optimization of the cellobiose utilization pathway, a pathway which is important in biofuel production. One strategy focused on optimizing enzyme combinations by creating a library of homologous proteins from the pathway. A second strategy investigated engineering all of the proteins in the pathway simultaneously. The improved pathway was assessed based on specific growth rate on cellobiose, with the final mutant exhibiting a 42% increase over the wild-type pathway. Metabolite analysis of the engineered pathway presented a 54% increase in cellobiose consumption (1.68 to 2.82 g cellobiose/(L•h)) and a 74% increase in ethanol productivity (0.59 to 1.03 g ethanol/(L•h)). The second half of the thesis was focused on creating a biofuel molecule with more diverse applications than the commonly used bioethanol biofuel. A new pathway for biodiesel production was investigated, using a heterologous fatty acid synthesis pathway, which would provide a completely orthologous route for biodiesel production. In this strategy, the endogenous fatty acid flux would not be redirected from cellular metabolism. Through heterologous expression of a Type-I fatty acid synthase, the total production of fatty acid ethyl esters was increased 6.3-fold, from 1670 µg FAEE/ g CDW to 10,498 µg FAEE/ g CDW. The final work in the thesis surveyed three potential high throughput screening methods to subject the biodiesel production pathway to the optimization strategies developed earlier in the thesis.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Chemical Engineering

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2014

Author dc:creator

• Eriksen, Dawn

Contributors dc:contributor

• Zhao, Huimin
• Harley, Brendan A.
• Schroeder, Charles M.
• Cann, Isaac K.

• combinatorial libraries
• pathway engineering
• biofuels
• microbial chemical production

Statement dc:rights

• Copyright 2014 Dawn Eriksen

Language dc:language

en

Handle dc:identifier

http://hdl.handle.net/2142/50368

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/50368