| Microorganisms can be used to produce a variety of chemicals such as drugs, enzymes, and fuels from different sugars. Traditionally, these processes have involved a single feedstock, most often glucose. More recently, significant effort has been devoted towards developing processes that directly use plant-based material as the feedstock. One challenge presented by the use of plant-based material is it is comprised of multiple sugars, each with a unique biochemistry. In particular, can these microorganisms process mixtures of sugars and, if not, is it possible to engineer them so that they can efficiently complete this task?;In this work, I investigate how Escherichia coli ( E. coli), a common industrial microorganism, uptakes and then metabolizes mixtures of hexose and pentose sugars, the main constituents in plant-based material. My specific focus was on understanding how the biochemical pathways for processing pentose sugars, such as arabinose and xylose, are regulated. In particular, E. coli will only activate a specific pathway if the target sugar is present. Furthermore, different pathways can interfere with one another in such a way that the processing of one sugar prevents that of another.;Using both genetic and analytical approaches, I discovered that E. coli will not simultaneously metabolize mixtures of hexose and pentose sugars. Instead, E. coli will sequentially process them based on their energy content. From an industrial standpoint, this hierarchy means that the conversion of these sugar mixtures will be inefficient and necessitate complex processing schemes. In order to understand the mechanism of how E. coli sequentially utilizes these sugars, I systematically removed the various steps in the hexose and pentose metabolic pathways, both individually and in combination. The results from these experiments allowed us to conclude that the key bottleneck is due to interference due to AraC, the arabinose protein that regulates transport and metabolism of the sugar.;Collectively, the results from my research have identified that E. coli employs a complex cellular control system in order to selectively process individual sugars. By identifying the mechanism for this control, I have identified specific genetic targets for subsequent metabolic engineering. This discovery will enable the construction of E. coli strains capable of simultaneously and efficiently processing mixtures of hexose and pentose sugars. These strains will likely have application in biotechnology and also potentially in the production of cellulosic biofuels. |
