A new metabolic engineering technique could reduce the cost and environmental impact of producing liquid biofuels and biochemicals, as alternatives to petroleum-based products.
In a new paper published in the journal Science, researchers at MIT and the Cambridge startup Novogy describe a new technique that gives producer microbes the upper hand against unwanted invaders, eliminating the need for such expensive and potentially harmful sterilisation methods.
The researchers engineered microbes, such as Escherichia coli, with the ability to extract nitrogen and phosphorous — two vital nutrients needed for growth — from unconventional sources that could be added to the fermentation vessels, according to Gregory Stephanopoulos, the Willard Henry Dow Professor of Chemical Engineering and Biotechnology at MIT, and Joe Shaw, Senior Director of Research and Development at Novogy, who led the research.
As the engineered strains only possess this advantage when they are fed these unconventional chemicals, the chances of them escaping and growing in an uncontrolled manner outside of the plant in a natural environment are extremely low.
“We created microbes that can utilise some xenobiotic compounds that contain nitrogen, such as melamine,” said Stephanopoulos. Melamine is a xenobiotic, or artificial, chemical that contains 67% nitrogen by weight.
Conventional biofermentation refineries typically use ammonium to supply microbes with a source of nitrogen. But contaminating organisms, such as Lactobacilli, can also extract nitrogen from ammonium, allowing them to grow and compete with the producer microorganisms.
In contrast, these organisms do not have the genetic pathways needed to utilise melamine as a nitrogen source, said Stephanopoulos. “They need that special pathway to be able to utilise melamine, and if they don’t have it they cannot incorporate nitrogen, so they cannot grow.”
The researchers engineered E. coli with a synthetic six-step pathway that allows it to express enzymes needed to convert melamine to ammonia and carbon dioxide, in a strategy they have dubbed ROBUST (Robust Operation By Utilisation of Substrate Technology).
When they experimented with a mixed culture of the engineered E. coli strain and a naturally occurring strain, they found the engineered type rapidly outcompeted the control, when fed on melamine.
They then investigated engineering the yeast Saccharomyces cerevisiae to express a gene that allowed it to convert the nitrile-containing chemical cyanamide into urea, from which it could obtain nitrogen.
The engineered strain was then able to grow with cyanamide as its only nitrogen source. Finally, the researchers engineered both S. cerevisiae and the yeast Yarrowia lipolytica to use potassium phosphite as a source of phosphorous.
Like the engineered E. coli strain, both the engineered yeasts were able to outcompete naturally occurring strains when fed on these chemicals.
“So by engineering the strains to make them capable of utilising these unconventional sources of phosphorous and nitrogen, we give them an advantage that allows them to outcompete any other microbes that may invade the fermenter without sterilisation,” said Stephanopoulos.
The microbes were tested successfully on a variety of biomass feedstocks, including corn mash, cellulosic hydrolysate, and sugar cane, where they demonstrated no loss of productivity when compared to naturally occurring strains.
The paper provides a novel approach to allow companies to select for their productive microbes and select against contaminants, according to Jeff Lievense, a senior engineering fellow at the San Diego-based biotechnology company Genomatica who was not involved in the research.
“In theory you could operate a fermentation plant with much less expensive equipment and lower associated operating costs,” said Lievense. “I would say you could cut the capital and capital-related costs (of fermentation) in half, and for very large volume chemicals, that kind of saving is very significant."
The ROBUST strategy is now ready for industrial evaluation, Shaw says. The technique was developed with Novogy researchers, who have tested the engineered strains at laboratory scale and trials with 1000 l fermentation vessels, and with Felix Lam of the MIT Whitehead Institute for Biomedical Research, who led the cellulosic hydrosylate testing.
Novogy now hopes to use the technology in its own advanced biofuel and biochemical production, and is also interested in licensing it for use by other manufacturers, Shaw said.
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Adapted from news report by Rosalie Starling
Read the article online at: https://www.hydrocarbonengineering.com/clean-fuels/08082016/mit-new-approach-could-lower-costs-for-biofuel-production-3874/