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The effect of pyruvate on the metabolic regulation of nitrogenase activity in Rhodospirillum rubrum with darkness as switch-off effector
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
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2010 (English)In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080Article in journal (Refereed) Submitted
Abstract [en]

Rhodospirillum rubrum, a photosynthetic diazotroph, is able to regulate nitrogenase activity in response to environmental factors such as ammonium ions or darkness – the so-called switch-off effect. This is due to reversible modification of the Fe-protein one of the two components of nitrogenase. The signal transduction pathway(s) in this regulatory mechanism is not fully understood, especially not in the response to darkness. We have previously shown that the switch-off response and metabolic state differ between cells grown with dinitrogen or glutamate as nitrogen source, although both represent poor nitrogen sources. In this study we show that addition of pyruvate to cultures grown with glutamate as nitrogen source will lead to a switch-off response that is similar to that in cultures grown with dinitrogen. The effects are related to PII protein uridylylation and GS activity. We also show that pyruvate induces de novo protein synthesis and that pyruvate formate-lyase activity is required for activity in the dark.

Place, publisher, year, edition, pages
2010.
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
URN: urn:nbn:se:su:diva-42159OAI: oai:DiVA.org:su-42159DiVA: diva2:351887
Available from: 2010-09-16 Created: 2010-08-18 Last updated: 2017-12-12Bibliographically approved
In thesis
1. Regulation of nitrogen fixation in Rhodospirillum rubrum: Through proteomics and beyond
Open this publication in new window or tab >>Regulation of nitrogen fixation in Rhodospirillum rubrum: Through proteomics and beyond
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Adaptability is one of the reasons for the success of bacteria, allowing them to survive in conditions where no other organisms would be able to thrive. Nitrogen deficiency, for example, can be a limiting factor for the growth of micro-organisms, as this element is an essential part of almost all types of biomolecules. As such, some bacteria have evolved specific mechanisms to overcome nitrogen limitation. Nitrogen fixing bacteria, or diazotrophs, use a specific enzyme complex, nitrogenase, in order to harness this element from the enormous reservoir that is the Earth’s atmosphere. However, nitrogen fixation is a demanding process for the cells, requiring vast amounts of energy and tight regulation.

In this thesis we explore the mechanisms regulating nitrogen fixation in Rhodospirillum rubrum, a purple non-sulphur photosynthetic bacterium. Using proteomics tools, we show how the regulation of both the nitrogen and carbon fixation processes is interconnected, possibly in order to maintain the intracellular redox balance. Using a new detergent molecule, we also demonstrate how nitrogen availability affects the chromatophore membrane proteome.

Our studies have revealed the crucial role of the cellular pool of 2-oxoglutarate (2OG) for adequate signaling through the PII proteins and the effects resulting from artificially manipulating this metabolite’s concentration. In R. rubrum nitrogenase is also subjected to post-translational control (the “switch-off” effect) and this work shows for the first time that the enzyme modifying nitrogenase (Dinitrogenase Reductase ADP-ribsosyl Transferase or DRAT) forms a complex with the PII protein GlnB. This complex allows DRAT activation and its formation – and, therefore, DRAT activity – is regulated by binding of ADP:ATP and 2OG to GlnB.

Upon light withdrawal, nitrogenase activity anaerobically in the dark is also here demonstrated to be dependent on the activity of the pathway starting in pyruvate formate-lyase and we show how different nitrogen sources influence the switch-off response. This response can, in some conditions, be modified by addition of pyruvate and we have studied how this metabolite influences nitrogenase activity and switch-off regulation.

This study allows a better understanding of the underlying processes controlling the metabolic routes in R. rubrum and also provides new insights into regulation of enzyme activity, paving the road for the complete establishment of the mechanisms regulating nitrogenase switch-off.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2010. 71 p.
Keyword
Rhodospirillum rubrum, nitrogen fixation, redox balance, switch-off, DRAT
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:su:diva-42101 (URN)978-91-7447-125-0 (ISBN)
Public defence
2010-10-08, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: In press. Paper 3: Submitted. Paper 4: Manuscript. Paper 5: Submitted.

Available from: 2010-09-16 Created: 2010-08-16 Last updated: 2012-10-03Bibliographically approved

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