Leigh Lab : Research
J o h n    L e i g h    L a b

University of Washington Department of Microbiology
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The Lab

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John Leigh, Ph.D.

Research : Overview

Our lab studies the species Methanococcus maripaludis, a member of the methanogenic Archaea. Our fascination with this organism stems in part from its representation of the third branch of life. As microorganisms that differ from the Bacteria and the Eucaryotes, the Archaea are ripe for discovery. Within the Archaea, the methanogens are important anaerobes in a variety of habitats. Methanococcus maripaludis is a superb model because of its excellent laboratory growth behavior, a robust set of genetic tools, and a complete genome sequence.

Here are the projects going on in the lab:

Electron flow and energy conservation in methanogens

We are working to understand the pathways of electron flow and the mechanisms of energy conservation in methanogens that lead to ATP synthesis. Using a genetic approach, we demonstrated support for the role of electron bifurcation as an energy-coupling mechanism, making the pathway of methanogenesis cyclical. We also showed the role of energy-converting hydrogenase Eha in providing an anaplerotic supply of electrons.

Engineering Methanococcus maripaludis for production of biofuels

Methanococcus maripaludis is an ideal host for engineering new metabolic pathways. As part of an ARPA-E funded collaboration, we are cloning genes into Methanococcus maripaludis that should enable it to produce methanol. In addition, we are interested in the production of hydrogen, and in the use of alternative substrates for production of methane. As part of this effort, we are collaborating with the laboratory of Nathan Price to develop a genome-scale metabolic flux model of Methanococcus maripaludis.

Nitrogen regulation

A wide variety of organisms sense cellular 2-oxoglutarate levels as an indicator of nitrogen starvation. Methanococcus maripaludis is no exception to this rule, as it responds to 2-oxoglutarate to regulate a variety of nitrogen assimilation functions, including nitrogen fixation. However, the components of the regulatory machinery have proven to be novel. We discovered the repressor NrpR and showed that it binds to the promoter regions of nitrogen-regulated genes, inhibiting transcription. 2-oxoglutarate acts as the inducer, preventing NrpR from binding to the DNA.

Another novel regulator is NifI. This protein is a relative of PII proteins, which measure 2-oxoglutarate levels and interact with a variety of other proteins that regulate nitrogen assimilation functions. NifI binds directly to nitrogenase and prevents its interaction with nitrogenase reductase. We are working to determine the structural basis for the function of NifI. So far we know that it undergoes interesting quaternary changes. When 2-oxoglutarate is low NifI is a hexamer and it presumably binds as a hexamer to nitrogenase. When 2-oxoglutarate is high, NifI appears to change into a dodecamer and does not bind nitrogenase.

We are also interested in methanogenic Archaea in the context of Astrobiology. Methanogenesis probably evolved early on earth and is a likely metabolism of any life that might exist on Mars or Europa. Members of the lab participate in the University of Washington's Astrobiology Program.

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