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Great Value in Industry: Converting Escherichia coli to a Synthetic Methylotroph Growing Solely on Methanol

Methanol is rich in electrons and can be derived from methane or carbon dioxide. It is a potential renewable one-carbon (C1) feedstock for microorganisms. The ribose monophosphate (RuMP) cycle used by methylotrophs to assimilate methanol is different from the typical sugar metabolism by only three enzymes (methanol dehydrogenase [Medh], hexose hexaphosphate synthase [Hps] and 6-phosphate-3-hexose isomerase [Phi]). However, it is still challenging to turn a non-methylotrophic organism to a synthetic methylotroph that grows to a high cell density.

The absorption of one-carbon (C1) compound by microorganisms has become a promising method to mitigate climate change. Among all C1 compounds, methanol is the liquid form with the most electrons and can be derived from the greenhouse gases (methane) and CO₂. Compared with those gaseous C1 compounds, methanol also avoids diffusion. In addition, methanol is already an industrial raw material, so it is potentially attractive as a substrate for bioconversion.

The natural methanol utilization and conversion pathways in methylotroph (such as Bacillus methanolicus) have been well characterized. Organisms usually use the ribose monophosphate (RuMP) cycle or serine pathway for methanol assimilation. Moreover, except for the three different enzymes mentioned above, the enzymes involved in the RuMP cycle overlap with those related to typical sugar metabolism. Therefore, for the benefit of science and industry, people have made great efforts to transform glycoheterotrophs into methylotrophs by overexpression of these three enzymes. If achieved, methanol can be used as an alternative non-food carbon source for common industrial microorganisms in the industrial biotechnology field.

Although the initial success has been achieved in the assimilation of methanol by engineering glycoheterotrophs, it has not been possible to convert such heterotrophs into methyl homotrophs that effectively use methanol as the only carbon and energy source. Obviously, the successful expression of the three heterologous genes is not enough to convert E. coli into a methylotroph.

Figure 1 Synthetic Methylotroph- E. Coli SM1 (Frederic Y.-H. Chen, et al. 2020)

Recently, a study published in Cell utilized metabolic robustness criteria to reprogram Escherichia coli (E.coli). Subsequently, laboratory evolution was carried out to establish a strain that can effectively use methanol as the sole carbon source. This synthetic methylotroph alleviated a so far uncharacterized hurdle, DNA-protein crosslinking (DPC), by insertion sequence (IS)-mediated copy number variations (CNVs) and balanced the metabolic flux by mutations. This synthetic methylotroph can grow at a rate comparable to that of natural methylotrophs in a wide range of methanol concentrations. This strain can explain genome editing and evolution for changes of microbial tropism, and expand the scope of biological C1 conversion.

Reference:

• Frederic Y.-H. Chen, et al. Converting Escherichia coli to a Synthetic Methylotroph Growing Solely on Methanol. Cell. 2020.