Date of Award
Master of Science (MS)
The advent of the industrial era was precipitated by the discovery of fossil fuels, and ushered in unprecedented changes for humanity included but not limited to the development of rapid transit and communications, improvements to food distribution and preservation, the mass production of goods, and a radical rearrangement of communities from relatively small enclaves to metropolises. With all the benefits, however, come considerable costs, especially to the global environment. Greenhouse gas emissions, built up over centuries of unregulated combustion, have precipitated a rate of global temperature change unparalleled in the 4.5 billion-year history of this planet. In order to preserve life on Earth, emissions must be dramatically reduced – first halved by 2030, and then diminished to a net-zero by 2050. One method to make these cuts is to explore renewable sources of energy and chemicals. One viable petroleum-alternative raw material is lignocellulosic biomass, the most abundant biopolymer in the world. Pretreatment of lignocellulose for depolymerization generates a range of compounds, including fermentable sugars which can be readily converted into alcohols for biofuel applications, and the recalcitrant aromatic portion, liginin. Due to costs associated with depolymerizing lignin and separating out the resulting mixture of heterogeneous, aromatic compounds, the lignin fraction is generally separated from the sugar-based cellulose and hemicellulose fractions and burned to supplement energy requirements of the biorefinery. Valorizing lignin into value-added compounds, however, not only helps to reduce greenhouse gas emissions, it also is a key factor in improving the economic viability of a biorefinery.
One of the more promising avenues for lignin valorization being explored is its use as a substrate for bioproduction of commodity chemicals by microbial cell factories. A confounding factor in this method is identifying microbes with the capability to tolerate and consume a wide range of toxic aromatic compounds, combined with the metabolic basis to become a bioproduction chassis. Rhodococcus opacus PD630 (hereafter PD630) is a Gram-positive, non-model soil bacterium isolated from soil outside a German gasworks plant and enriched on phenyldecane. Among its desirable traits are a diverse range of compatible substrates ranging from sugars to lignin-derived aromatics, the ability to accumulate more than half its dry cell weight in triacylglycerols (TAGs, a precursor for biofuels and other chemicals), a rapidly-developing genetic toolbox, and moderately fast growth. This thesis examines the development of PD630 as a platform for lignin valorization, focusing in particular on conditions promoting TAG accumulation. To this end, the research herein presents a method for mining transcriptomic datasets for regulatory genes which are part of the response network for particular environmental conditions. In brief, a regulatory element is identified which supports nitrogen-independent accumulation of TAG, particularly when phenol is present in the media. As TAG production is largely upregulated in response to nitrogen starvation, and nitrogen stress inhibits aromatic tolerance in PD630, these findings represent progress toward optimizing PD630 as a biocatalyst for lignin valorization. Additionally, they elucidate some of the network rearrangements necessary to support PD630 switching from a growth phenotype to TAG accumulation in a nitrogen-depleted environment.
Tae Seok Moon
Yinjie Tang Marcus Foston