Rutgers 2012 iGEM Team: Biofuels in Biology

Rutgers 2012 iGEM Team: Biofuels in Bacteria


The current fossil fuel-dependent economy drives a demand for sustainable energy resources. Although much effort has gone into the production of ethanol, other biofuels, such as butanol, are superior. Butanol has a higher energy content, is less volatile, and is safer to use than ethanol.

To develop strains of bacteria that produce high levels of 1-butanol we have introduced the genes coding for a biochemical pathway from Clostridium acetobutylicum into a mutant E. coli strain that produces a high level of NADH.

The combination of these chemical pathways is predicted to increase the level of butanol production.

Biobutanol: Origins and Prospects

Genetically modified biological systems can provide direct industrial approaches to the production of commodity chemicals.  The ability to manipulate chemical pathways with the tools of synthetic biology has opened new doors in the renewable energy industry.  This year, the Rutgers iGEM team has engineered a bacterial strain that can produce 1-butanol, a highly efficient biofuel that is able to generate up to 95% the energy produced by the combustion of gasoline.

Butanol is a substitute fuel for gasoline that can be produced by ABE (acetone, butanol, ethanal) fermentation, which is the anaerobic conversion of carbohydrates into alcohol-based fuels.

The anaerobic conversion of carbohydrates by strains of Clostridium into acetone, butanol and ethanol (not a sentence). However, cost issues, the relatively low-yield, and slow fermentation processes, as well as problems caused by end product inhibition and phage infections, meant that butanol generated by ABE fermentation could not compete on a commercial scale with butanol produced synthetically.

There is now increasing interest in use of biobutanol as a transport fuel. Butanol produced from bacteria, or biobutanol can be made from a variety of sources including biomass, or simple sugars such as glucose.

The fermentation of sugars into butanol was first completed by Dr. Chiam Weizmann.  

Dr. Weizmann worked with a strain of bacteria known as Clostridium acetobutylicum.  His original experiment was set up to ferment starch into acetone, unexpectedly, Dr. Weizmann made twice the amount of an unwanted side product-butanol.  

Biobutanol is highly similar to gasoline, and remains highly advantageous over other alcohol based fuels.

A simple 85% butanol 15% gasoline mixture can be used in today’s fuel pipelines and internal combustion engines without any modifications to the current fuel system. 

  • Butanol is hydrophobic, and has lower tendency to mix with water molecules that may cause fuel contamination.

  • Ethanol poses the issue of water contamination and due to its corrosive properties, causing damage to the fuel pipelines.  

Not only is butanol better than other alcohol-based fuels in terms of molecular properties, but it also possess a higher energy content and air-fuel ratio.

Butanol Production in E. Coli

We have focused on implementing a 1-butanol producing biochemical pathway, initially found in Clostridium, into the very well known model organism E. coli.

The natural production of 1-butanol in Clostridium has been introduced to a variety of species such as Saccharomyces cerevisiae, Lactobacillus brevis, Pseudomonas putida, and Bacillus subtilis. All of these organisms produced a low-titer amount of 1-butanol. The introduction of this biochemical pathway into E. coli has led to the production of more traceable amounts of 1-butanol.  

We have designed a highly intricate system of promoters and operators organized to break down glucose, a complex carbohydrate, into a series of intermediary molecules before 1-butanol is obtained.

The Clostridium derived butanol pathway features a network of eight genes and three promotors:

  • fdh
  • ter
  • adhE2
  • crt
  • phaA2
  • araC
  • hbd
  • aceE
  • aceF
  • lpd
  • arabinose BAD promoter
  • trc promoter
  • lac promoter

The combination of these chemical pathways is predicted to increase the level of butanol production.

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Parts Submission

RUiGEM2012 -- Parts sent to the Registry