Reversing methanogenesis to capture methane for liquid biofuel precursors

Abstract

BackgroundEnergy from remote methane reserves is transformative; however, unintended release of this potent greenhouse gas makes it imperative to convert methane efficiently into more readily transported biofuels. No pure microbial culture that grows on methane anaerobically has been isolated, despite that methane capture through anaerobic processes is more efficient than aerobic ones.ResultsHere we engineered the archaeal methanogen Methanosarcina acetivorans to grow anaerobically on methane as a pure culture and to convert methane into the biofuel precursor acetate. To capture methane, we cloned the enzyme methyl-coenzyme M reductase (Mcr) from an unculturable organism, anaerobic methanotrophic archaeal population 1 (ANME-1) from a Black Sea mat, into M. acetivorans to effectively run methanogenesis in reverse. Starting with low-density inocula, M. acetivorans cells producing ANME-1 Mcr consumed up to 9 ± 1 % of methane (corresponding to 109 ± 12 µmol of methane) after 6 weeks of anaerobic growth on methane and utilized 10 mM FeCl3 as an electron acceptor. Accordingly, increases in cell density and total protein were observed as cells grew on methane in a biofilm on solid FeCl3. When incubated on methane for 5 days, high-densities of ANME-1 Mcr-producing M. acetivorans cells consumed 15 ± 2 % methane (corresponding to 143 ± 16 µmol of methane), and produced 10.3 ± 0.8 mM acetate (corresponding to 52 ± 4 µmol of acetate). We further confirmed the growth on methane and acetate production using 13C isotopic labeling of methane and bicarbonate coupled with nuclear magnetic resonance and gas chromatography/mass spectroscopy, as well as RNA sequencing.ConclusionsWe anticipate that our metabolically-engineered strain will provide insights into how methane is cycled in the environment by Archaea as well as will possibly be utilized to convert remote sources of methane into more easily transported biofuels via acetate.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-015-0397-z) contains supplementary material, which is available to authorized users.

Metabolic manipulation of methanogens for methane machinations

“…Global amounts of shale gas total 7300 trillion cubic feet (U.S. Energy Information Administration, 2013) and its major constituent is methane. This remarkable availability of methane is now driving synthetic biology, and an exciting prediction is that methane will be harnessed for biotechnological applications using not the traditional workhorse E. coli or aerobic methanotrophs , but instead, using archaeal strains, specifically methanogens, in anaerobic fermentations based on biosynthetic pathways such as that recently shown to convert methane to the biotechnological building block acetate (Soo et al ., 2016). …”

“…The first process used to capture methane anaerobically for biotechnology applications (Soo et al ., 2016) is based on the natural process of anaerobic methane oxidation (AOM), which efficiently captures up to 300 Tg of methane per year to limit global methane emissions (Knittel and Boetius, 2009). AOM occurs in natural consortia consisting of anaerobic methanotrophic archaeal populations and syntrophic bacteria.…”

“…In effect, the first anaerobic organism that grows on methane as a pure culture was created. Remarkably, the engineered strain grows as a biofilm on solid ferric chloride which was reduced by the electrons generated by growth on methane (Soo et al ., 2016). Carbonate in the medium and methane in the headspace were converted into the two carbons of acetate as shown by 13 C labelling of both substrates (Soo et al ., 2016).…”

“…Remarkably, the engineered strain grows as a biofilm on solid ferric chloride which was reduced by the electrons generated by growth on methane (Soo et al ., 2016). Carbonate in the medium and methane in the headspace were converted into the two carbons of acetate as shown by 13 C labelling of both substrates (Soo et al ., 2016). These results in effect put an end to the decade's old debate about whether methane can be fixed by running methanogenesis in reverse (Knittel and Boetius, 2009).…”

“…Although not an end in itself, the acetate produced from methane by reversing methanogenesis (Soo et al ., 2016) is a building block for many products. For example, via additional metabolic engineering and production of active methanotrophic Mcr, we have now converted methane into the pure stereoisomer L‐ lactate which may be used in cosmetics, foods and pharmaceuticals.…”