(143f) Making Escherichia coli Grow on Methanol

The quest for using methanol as a substrate to produce useful metabolites such as commodity chemicals, chemical precursors, biopolymers, neutraceuticals and, more recently, biofuels, goes back to more than 40 years. The concept originated from the development of processes to produce single-cell protein (SCP) from methanol or methane using native methylotrophs. While technologically successful, SCP did not succeed in the marketplace, but led to many technological innovations in bioreactor design. The more recent increasing availability and lower prices of natural gas is resulting in increased methanol supplies, and has thus renewed the interest in using methanol as a substrate or co-substrate with sugars. The latter derives from the fact that methanol is considerably more reduced than most sugars and thus has to potential to enable the biological production of metabolites with significantly increased yields. The biology of natural methylotrophs has generated a large body of knowledge, but little of that has translated into biotechnological applications despite the sustained research enterprise. The last 10-15 years has also seen efforts to engineer other organisms to use methanol for growth, but without significant success yet. E. coli is an obvious platform for synthetic methylotrophy. We will discuss the native biology of methylotrophy and how it can be engaged to develop superior methylotrophic strains using synthetic biology and metabolic engineering. In principle, E. coli and most other organisms need only 3 genes/enzymes to be able to use methanol as a substrate, and tried though it may have been, an E. coli or other non-methylotrophic organism that can effectively grow on methanol as a single substrate has not been reported. There are several reasons for that, and many barriers to be overcome: the choice of desirable enzyme co-factors, thermodynamic roadblocks and, most importantly, regulatory constraints that make the quest for synthetic methylotrophy an exciting challenge. We will discuss how we systematically dissected the issues of synthetic methylotrophy to arrive to problem resolution. SUPPORTED by the US DOE ARPA-E agency through contract no. DE-AR0000432.