Creating synthetic metabolic pathways for carbon dioxide fixation could reduce atmospheric carbon dioxide levels while providing a carbon-neutral alternative to conventional chemical manufacturing processes for pharmaceuticals and active ingredients. For example, a recent study in Nature Communications illustrates a method of transforming carbon dioxide into a beneficial substance for the biochemical industry through the use of formic acid.

Considering the increasing greenhouse gas emissions, finding effective carbon capture methods, specifically those that sequester carbon dioxide from significant emission sources, is imperative. Although carbon dioxide assimilation has occurred naturally for millions of years, its capacity does not offset human-induced emissions.

Under the leadership of Tobias Erb at the Max Planck Institute for Terrestrial Microbiology, a team of researchers is leveraging nature's mechanisms to explore new carbon dioxide fixation methods. They have created a synthetic metabolic pathway that generates highly reactive formaldehyde from formic acid, a potential byproduct of artificial photosynthesis. Formaldehyde can be directly integrated into several metabolic pathways to produce other valuable substances without harmful effects. Like the natural one, this process necessitates two primary elements: Energy and carbon. The energy can be supplied by direct sunlight and electricity, such as from solar panels.

Within the value-added chain, the carbon source can vary. Carbon dioxide is not the sole option; all monocarbons (C1 building blocks), such as carbon monoxide, formic acid, formaldehyde, methanol, and methane, can be considered. However, most of these substances pose a toxic threat to living organisms (carbon monoxide, formaldehyde, methanol) or the environment (methane as a greenhouse gas). Only formic acid, when neutralized to its base formate, is tolerated by many microorganisms in high concentrations.

"Formic acid is an exceptionally promising carbon source," emphasizes Maren Nattermann, the study's lead author. Yet, she adds that the transformation of formic acid to formaldehyde is quite energy-demanding. The challenge lies in the fact that the salt of formic acid, formate, does not easily convert into formaldehyde. Overcoming the significant chemical hurdle between the two molecules requires biochemical energy, in the form of ATP, before the actual reaction can be performed.

The researchers aimed to discover a more efficient method. After all, the less energy required to integrate carbon into metabolism, the more energy is available for promoting growth or production. Finding such a pathway in nature, however, is not possible. Tobias Erb notes that creativity is required to uncover so-called promiscuous enzymes with multiple functions. Still, identifying potential enzymes is just the starting point, as some reactions are so slow they can be counted individually. Natural reactions can be thousands of times quicker. This is where synthetic biochemistry steps in, says Maren Nattermann, adding, "If you understand an enzyme's structure and mechanism, you know how to intervene."

“We developed a two-enzyme route in which formate is activated to formyl phosphate and subsequently reduced to formaldehyde. Exploiting the promiscuity of acetate kinase and N-acetyl-γ-glutamyl phosphate reductase, we demonstrate this phosphate (Pi)-based route in vitro and in vivo,” the authors wrote “We further engineered a formyl phosphate reductase variant with improved formyl phosphate conversion in vivo by suppressing cross-talk with native metabolism and interface the Pi route with a recently developed formaldehyde assimilation pathway to enable C2 compound formation from formate as the sole carbon source in Escherichia coli. The Pi route, therefore, offers a potent tool in expanding the landscape of synthetic formate assimilation.”

The process of enzyme optimization involved several strategies, including specific exchange of building blocks, generating random mutations, and selecting for capabilities. "Both formate and formaldehyde are ideal because they can cross cell walls. Therefore, we introduce formate into the culture medium of cells producing our enzymes, and within a few hours, the resulting formaldehyde is converted into a non-toxic yellow dye," Maren Nattermann explains.

This achievement was accelerated with the help of high-throughput methods, a process facilitated through a partnership with industrial partner Festo, based in Esslingen, Germany. "After testing approximately 4000 variants, we witnessed a fourfold increase in production," says Maren Nattermann. "We have established the groundwork for the model microbe Escherichia coli to grow on formic acid. However, our cells currently only produce formaldehyde and cannot convert it further."

Collaborating with Sebastian Wenk at the Max Planck Institute of Molecular Plant Physiology, the team is developing a strain that can assimilate and introduce the intermediates into the central metabolism. Concurrently, the team is researching the electrochemical conversion of carbon dioxide to formic acid at the Max Planck Institute for Chemical Energy Conversion, led by Walter Leitner. The ultimate aim is to create a comprehensive platform to transform carbon dioxide into products like insulin or biodiesel through an electrobiochemical process.

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