Team:Aalto-Helsinki

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Team:WashU StLouis/Header

Washington University

Engineering Ntrogen Fixation

Washington University in St. Louis
Engineering Nitrogen Fixation

Project Description

Some cyanobacteria fix nitrogen for nutritional needs, while most organisms can only acquire it from the food it consumes. Synthetic biology allows us to transfer this ability to fix nitrogen to a heterologous host that has many genetic tools, Escherichia coli, so that we can learn how to give single cell organisms, and eventually chloroplasts the ability to create their own nitrogen fertilizer.

Diazotrophic (organisms that fix nitrogen) cyanobacteria such Nostoc Punctiforme or Anabaena use heterocysts (specialized nitrogen fixing cells) to create a mini-anaerobic environment to aid nitrogen fixation. However, Cyanothece 51142, a non-heterocyst, fixes nitrogen in the same cell as photosynthesis by relying on a circadian metabolic process, when there is less oxygen byproduct from photosynthesis. This process is both fascinating and necessary since the key enzyme in nitrogen fixation, nitrogenase, is poisoned by oxygen. Our goal this summer is to engineer the regulation of the proteins necessary for nitrogen fixation so that they are highly repressed when activated by broad spectrum light (such as the sun), and are highly active when there is no light around, mimicking the cycle where photosynthesis occurs during the day and nitrogen fixation occurs at night.

Our pitch video.

How does our project work?

Our team worked this summer on two projects towards a common purpose. Richard and Caroline worked in the Pakrasi lab under Dr. Pakrasi, with Andrew Ng, Bert Berla, and Deng Liu as advisors on the getting nitrogen fixation working in E. coli. Benjamin and Jeffrey worked in the Moon lab under Dr. Moon, with Cheryl Immethun as an advisor, and created a repressor system to turn off transcription of a reporter protein in the light.

What did we accomplish?

There are two key components of our project:

Richard and Caroline have been working in the Pakrasi lab, and have been using genes from cyanobacteria to get nitrogen fixation working in E. coli . They are testing nitrogen fixation by running acetylene reduction assays and designing experiments to test the optimal criteria (ie. E. coli strains, temperature, pH, nitrogen source) to get maximum results. For more info, visit the nitrogenase tab.

Who will our project help?

Our project is the first step of a much larger, much more complex endeavor. Nitrogen overabundance and nitrogen depletion are simultaneously big stumbling blocks in modern agriculture. The solution to both of these problems would be to endow plants themselves with the ability to fix nitrogen so that they could autonomously supply their own nitrogen for proteins, DNA, etc. We are taking the first step towards this ambitious goal by studying how the genes for nitrogen fixation from cyanobacteria work in different environments and constructing an artificial transcriptional system. We are currently working in E. coli because it is easy to engineer, but the next step would be to move into a cyanobacteria more closely related to chloroplasts. We hope that by making these initial steps that we may be helping to pave the way for future research that may put an end to world hunger.

Accomplishments

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