Glycolysis is currently used in biorefineries to convert sugars derived from plant biomass into biofuels, but the loss of two carbon atoms for every six that are input is seen as a major gap in the efficiency of the process. The wasted CO₂ leads to a significant decrease in carbon yield, the researchers observed, which results in a major impact on the overall economy of biorefinery and the carbon efficiency of cell growth. However, re-fixing the lost CO₂ would incur energetic and kinetic costs.

While it is theoretically possible to split sugars or sugar phosphates into stoichiometric amounts of acetyl-CoA in a carbon- and redox-neutral manner, resulting in maximal yields, no such pathways are known to exist in nature.

The UCLA research team addressed that lack, developing the synthetic glycolytic pathway which converts all six glucose carbon atoms into three molecules of acetyl-CoA without losing any as carbon dioxide.

Glycolysis, or its variations, is a fundamental metabolic pathway in life that functions in almost all organisms to decompose external or intracellular sugars. The pathway involves the partial oxidation and splitting of sugars to pyruvate, which in turn is decarboxylated to produce acetyl-coenzyme A (CoA) for various biosynthetic purposes. The decarboxylation of pyruvate loses a carbon equivalent, and limits the theoretical carbon yield to only two moles of two-carbon (C2) metabolites per mole of hexose. This native route is a major source of carbon loss in biorefining and microbial carbon metabolism.

Here we design and construct a non-oxidative, cyclic pathway that allows the production of stoichiometric amounts of C2 metabolites from hexose, pentose and triose phosphates without carbon loss. We tested this pathway, termed non-oxidative glycolysis (NOG), in vitro and in vivo in Escherichia coli. NOG enables complete carbon conservation in sugar catabolism to acetyl-CoA, and can be used in conjunction with CO₂ fixation and other one-carbon (C1) assimilation pathways to achieve a 100% carbon yield to desirable fuels and chemicals.

—Bogorad et al.

Dr. Liao, the principle investigator, is UCLA’s Ralph M. Parsons Foundation Professor of Chemical Engineering and chair of the chemical and biomolecular engineering department. Igor Bogorad, a graduate student in Liao’s laboratory, is the lead author.

This pathway solved one of the most significant limitations in biofuel production and biorefining: losing one-third of carbon from carbohydrate raw materials. This limitation was previously thought to be insurmountable because of the way glycolysis evolved.

—James Liao

This synthetic pathway uses enzymes found in several distinct pathways in nature.

The team first tested and confirmed that the new pathway worked in vitro. Then, they genetically engineered E. coli bacteria to use the synthetic pathway and demonstrated complete carbon conservation. The resulting acetyl-CoA molecules can be used to produce a desired chemical with higher carbon efficiency. The researchers dubbed their new hybrid pathway non-oxidative glycolysis, or NOG.

This is a fundamentally new cycle. We rerouted the most central metabolic pathway and found a way to increase the production of acetyl-CoA. Instead of losing carbon atoms to CO₂, you can now conserve them and improve your yields and produce even more product.

—Igor Bogorad

The researchers also noted that this new synthetic pathway could be used with many kinds of sugars, which in each case have different numbers of carbon atoms per molecule, and no carbon would be wasted.

For biorefining, a 50 percent improvement in yield would be a huge increase. NOG can be a nice platform with different sugars for a 100 percent conversion to acetyl-CoA. We envision that NOG will have wide-reaching applications and will open up many new possibilities because of the way we can conserve carbon.

—Igor Bogorad

The researchers also suggest this new pathway could be used in biofuel production using photosynthetic microbes.

The paper’s other author is Tzu-Shyang Lin, who recently received a bachelor’s degree from UCLA in chemical engineering.

Resources

• Igor W. Bogorad, Tzu-Shyang Lin & James C. Liao (2013) Synthetic non-oxidative glycolysis enables complete carbon conservation. Nature doi: 10.1038/nature12575