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Researchers generate methane from CO2 in one light-driven step using engineered bacteria

Using an engineered strain of the phototropic bacterium Rhodopseudomonas palustris as a biocatalyst, a team from the University of Washington, Utah State University and Virginia Polytechnic Institute and State University have reduced carbon dioxide to methane in one enzymatic step.

The work demonstrates the feasibility of using microbes to generate hydrocarbons (i.e., CH4 in this case) from CO2 in one enzymatic step using light energy. A paper on their work is published in Proceedings of the National Academy of Sciences (PNAS).

An essential process for life and an important step in the biogeochemical nitrogen cycle is nitrogen fixation by nitrogenase, in which nitrogen gas (N2) is converted to ammonia (NH3).… Nitrogenase deprived of access to N2 but provided with a source of electrons produces H2 exclusively. Also, the ability of nitrogenase to carry out the multielectron reduction of an inert molecule is not limited to reduction of N2. This enzyme can also reduce other molecules with double and triple bonds, including carbon-containing compounds. Recently, we found that a remodeled nitrogenase with substitutions in two key amino acids near the FeMo cofactor is capable of reducing carbon dioxide (CO2) to methane (CH4) in vitro. This enzyme did not retain its ability to reduce N2 but was active in H2 production. It was unclear if the remodeled nitrogenase gene could confer to bacteria the ability to reduce CO2 to CH4.

Here, we describe a biocatalyst capable of generating the energy-rich hydrocarbon CH4 by reduction of CO2 using a remodeled nitrogenase. Development of this biocatalyst required selection of an appropriate microbial host … We reasoned that the anoxygenic photosynthetic bacterium Rhodopseudomonas palustris would be a good chassis for studying the remodeled nitrogenase in the context of a biological system, because it can fix nitrogen, and an activating mutation in nifA, encoding the transcription activator of nitrogenase genes, has been identified that can bypass the regulatory networks that repress nitrogenase. Also, R. palustris can generate the considerable amount of ATP needed for the activity of a remodeled nitrogenase from light by cyclic photophosphorylation.

—Fixen et al.
Metabolic route of CH4 production by an engineered strain of R. palustris expressing a remodeled nitrogenase. ATP is produced by cyclic photo-phosphorylation, in which electrons energized by light are cycled through a proton-pumping electron transport chain rather than transferred to a terminal electron acceptor. Electrons are generated by oxidation of (A) organic compounds or (B) inorganic compounds, such as thiosulfate. CO2 from bicarbonate or generated from acetate oxidation is converted by remodeled nitrogenase to CH4 and the CBB cycle to cell material. Fixen et al. Click to enlarge.

Rhodopseudomonas palustris is an area of emphasis for the lab group of Dr. Caroline Harwood, University of Washington Professor of Microbiology and Associate Vice-Provost for Research, and corresponding author of the PNAS paper.

R. palustris is a purple photosynthetic bacterium that lives in environments like the surface layers of water logged soils that tend to straddle oxic to anoxic transition zones. When native Rhodopseudomonas is exposed to atmospheric levels of oxygen, it oxidizes carbon compounds such as acetate for carbon and energy by aerobic respiration. When it is exposed to oxygen depleted (2-6% oxygen) or anaerobic conditions, R. palustris turns a characteristic deep purple color as it synthesizes the light absorbing pigments that it needs to carry out photosynthesis.

Under microaerobic or anaerobic growth conditions R. palustris can use nitrogen gas from the atmosphere as a sole nitrogen source for growth by the process of nitrogen fixation. Rhodopseudomonas generates hydrogen along with ammonium as a product of nitrogen fixation.

In the study, the team found that engineered R. palustris reduced CO2 to CH4 using a remodeled nitrogenase when the enzyme was expressed in a genetic background that allows for its constitutive production and cells were incubated in light.

They also found that R. palustris is a tractable system that allows control of CH4 production by manipulating the distribution of electrons and energy available to nitrogenase.

It’s a baby step, but it’s also a big step. Imagine the far-reaching benefits of large-scale capture of environmentally damaging byproducts from burning fossils fuels and converting them to alternative fuels using light, which is abundant and clean. To our knowledge, no other organism can achieve what this bacterium has done with a single enzyme.

—USU professor Lance Seefeldt

Its ability to reduce carbon dioxide at ambient temperature and pressure with a single enzyme and energy provided by light makes the methane-producing strain of R. palustris an excellent starting point to understand how a biological system can marshal resources to produce an energy-rich hydrocarbon in one enzymatic step.

—Fixen et al.

The team’s work is supported by a grant awarded through the U.S. Department of Energy Office of Science’s Energy Frontier Research Center program to the Center for Biological and Electron Transfer and Catalysis or “BETCy.”


  • Kathryn R. Fixen, Yanning Zheng, Derek F. Harris, Sudipta Shaw, Zhi-Yong Yang, Dennis R. Dean, Lance C. Seefeldt, and Caroline S. Harwood (2016) “Light-driven carbon dioxide reduction to methane by nitrogenase in a photosynthetic bacterium” PNAS doi: 10.1073/pnas.1611043113


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