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Expression of the PII-AmtB encoding operons in Rhodospirillum rubrum and studies of the functional role(s) of GlnB, GlnJ and AmtB1 in nitrogen metabolism
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 Botany.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

In Rhodospirillum rubrum and many other diazotrophs, PII proteins and the ammonium transport protein AmtB have been shown to play central roles in the regulation of nitrogen metabolism. In this report we have used Real-time RT-PCR to study the transcription of the genes encoding three PII proteins and the AmtB proteins in R. rubrum. We have generated amtB1 and amtB2 mutants and in the amtB1 mutant strains ammonium nitrogenase switch-off is lost although the rate of ammonium uptake is not affected compared to wild type.  In contrast darkness switch-off is unaffected. Most interestingly, we also show that the uridylylation status of GlnB is different from that of GlnJ under certain conditions in the amtB1 mutant strain, which is the first demonstration of physiological selectivity in PII modification in vivo, supporting the proposed different functions for these paralogs in the cell.  We suggest that the primary role of AmtB1 in this diazotroph, is as a central component together with GlnJ in signal transduction regulating nitrogen metabolism.

Research subject
URN: urn:nbn:se:su:diva-30812OAI: diva2:274155
Available from: 2009-10-27 Created: 2009-10-27 Last updated: 2010-01-12Bibliographically approved
In thesis
1. PII proteins as global regulators of bacterial nitrogen metabolism
Open this publication in new window or tab >>PII proteins as global regulators of bacterial nitrogen metabolism
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Nitrogen is an essential element to sustain life, being a component of most biological macromolecules. In spite of the abundance of gaseous N2, the availability of nitrogen compounds that can be readily used by most microorganisms is scarce and its production energetically demanding. Due to the central importance of nitrogen metabolism, most microorganisms evolved elaborate mechanisms to ensure efficient regulation, balancing substrate availability, product formation and energy expenditure.

In most bacteria, many archaea and some plants, the different aspects of nitrogen metabolism are coordinated by members of the PII family of signal transduction proteins, acting as fundamental molecular messengers controlling several cellular processes. In proteobacteria, including the nitrogen fixing organism Rhodospirillum rubrum, these proteins are involved in regulation at different levels: they regulate gene expression, modulating the activity of several transcription factors; they control the flux through the ammonium transport protein (AmtB); they influence the activity of key metabolic enzymes, e.g. glutamine synthetase (GS) and nitrogenase. The signal sensing and integration by these proteins is achieved in two different yet interdependent strategies: allosteric regulation (by the binding of metabolites like ATP, ADP, 2-oxoglutarate) and reversible post-translational modification. Signal integration likely results in different conformations of the proteins, influencing the direct protein-protein interaction with the cellular targets.

In the present work, using R. rubrum as a model organism, we have studied some aspects of the biochemistry of PII proteins in terms of regulatory interactions with the ammonium transport protein AmtB1 and the adenylyltransferase GlnE (involved in GS regulation). Additionally, we have investigated the post-translational modification of PII proteins, showing for the first time in vivo in addition in vitro selectivity in the modification of different PII proteins.

Our results contributed to elucidate several new aspects in the regulation by PII proteins and also strengthened the idea that these proteins act as global regulators in the context of bacterial nitrogen metabolism.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholms universitet, 2009. 68 p.
Rhodospirillum rubrum, nitrogen metabolism, signal transduction, PII proteins
National Category
Biochemistry and Molecular Biology
Research subject
urn:nbn:se:su:diva-30815 (URN)978-91-7155-963-0 (ISBN)
Public defence
2009-12-07, Magnélisalen, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
At the time of the doctoral defence the following papers were unpublished and had a status as follows: Paper 1: Manuscript. Paper 4: Manuscript.Available from: 2009-11-15 Created: 2009-10-27 Last updated: 2010-01-12Bibliographically approved

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Teixeira, Pedro FilipeNordlund, Stefan
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