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Journal Club:”An Engineered Microbial Platform for Direct Biofuel Production from Brown Macroalgae”

Cover of Science Issue 335.


It has been a while since we have done a journal club, and so today I thought I would write about an article I recently read. Adam J. Wargacki, who works in Yasuo Yoshikuni's group, recently published the above titled paper.

In light of problems associated with the use of fossil energy sources (cost, scarcity, environmental impact, and geopolitical considerations), the search for new energy sources is starting to become more important. The paper in discussion today proposes that a coupled system consisting of brown macroalgae and engineered bacteria could be used.

Of course there are more traditional biofuel feedstock sources such as corn and sugarcane. There a couple of hurdles such as the debate about “food versus fuel” and technical hurdles such as the degradation of lignocellulosic matter that need to be solved. We have previously discussed how microalgaecould get around the difficulties of corn and sugarcane in the production of bioefuels. At the current time, genetically manipulating these algae is technically difficult. So Bioarchitecture Labs is proposing the use of brown macroalgae (ak seaweed) as an industrial feedstock.


Why?

The group claims that macroalgae have a couple of advantages. For one, these macroalgae are grown in sea water which does not compete with rare sweet water resources. This also means that macroalgae are not cultivated in agricultural lands used for food production leaving the “food vs fuel” debate out. Lastly, brown macroalgae do not contain lignin which means that sugars required for the production of biofuels can easily be obtained by crushing the harvested biomass.

What did Bio Architecture Labs actually do?

Logo of Bio Architecture Labs
Of course there are challenges trying to use brown algae as a biofuel feedstock. The challenges consists of the fact that brown algae are rich in other sugars (mainly alginate, mannitol and glucan). The degradation of manitol and glucan by bacteria is associated with metabolic imbalances generating excess reducing equivalents. The group realized that the challenges associated with manitol break-down could be counteracted by also breaking down alginate which consumes two reducing equivalents. The problem: Most bacteria including E. coli which is a well-known model organism strain that has traditionally been used for other biotechnological applications do not have pathways to degrade alginate. And thus, most of the paper describes how Bio Architecture Labs was able to still use seaweed as a feedstrock.

The group first showed that E. coli transformed with plasmid containing an engineered alginate lyase was able to secrete and degrade alginate effectively. Next, the group successfully identified and incorporated a functional heterologous alginate transport and metabolic system into E. coli. The paper describes this step as the most challenging step because the only alginol transport system known until then consisted out of a complicated outer-membrane protein complex and inner-membrane ABC transporter system which was never successfully shown to be incorporated into E. coli. Instead, the group ended up trying to find a way around the challenge of incorporating that system by relying on homology searches of simpler transporter and symporter systems that have previously been successfully incorporated into E. coli. The group found a hypothetical match in a 30 kb fragment in the genome of Vibro spledidus. Bio Architecture Labs then created a fosmid (similar to cosmids but more stable) library and screened for fragments that would allow E. coli cells to grow on medium containing alginate oligomers. The research group identified three genes this way that were previously also found using the homology search strategy. To prove the necessity and functionality of these genes in alginate import and metabolism, each gene was individually knocked out and tested. Lastly, the group looked for further genes that could help in the import and metabolism of alginate by looking for other genes surrounding the initial 30 kb fragment and identified a couple of more candidates which the group further characterized.

The result: After several rounds of optimization on synthetic medium, E. coli containing the full alginate metabolism pathway were able to produce ethanol directly from Saccharina japonica at a final titer of ~ 4.7% which represents an approximate yield of 80% of maximum theoretical yield. This represents terrific efficiencies in the pathway.

So what's the significance?

Erik Stokstad noted in a commentary appearing in the same issue of Science that E. coli could be modified to produce other fuels and other valuable chemicals from the same feedstock source. As to major hurdles and sustainability, the commentary points out that to just replace 1 % of U.S. Gasoline supply would require growing seaweed in an area of over 11, 000 square kilometers. To put this into perspective, the US does not currently do any significant amount of seaweed farming, and due to the large area requirement the technology would obviously only be feasible in coastal regions. Still, in the search for alternative energy sources, there will be no silver bullet. Seaweed-derived biofuels can be one of many important sources. Or cautionary note, according to the commentary, a full-life cycle analysis is currently being performed right now so there is no final answer yet when it comes to sustainability aspects. My guess is that seaweed-derived biofuels can be sustainable if a number of stringent criteria are fulfilled. Of course, for the reader of this blog, the question of coupling several processes such as bioremediation with production of renewable chemicals is always an interesting aspect to further explore.

Source:

1. Adam J. Wargacki et. al. “An Engineered Microbial Platform for Direct Biofuel Production from Brown Macroalgae” Science 335, 308 (2012)
2. Erik Stoktad “Engineered Superbugs Boost Hopes Of Turning Seaweed Into Fuel”. Science 335, 273 (2012).

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