Metabolic regulation of nitrogen fixation in Rhodospirillum rubrum

University dissertation from Stockholm : Department of Biochemistry and Biophysics, Stockholm University

Abstract: Nitrogen, along with carbon, hydrogen and oxygen, is amongst the most abundant elements in all living cells. The capability to convert atmospheric dinitrogen to biologically usable nitrogen compounds is only found in some prokaryotes. Biological nitrogen fixation, the reduction of dinitrogen to ammonia, is the entry step into the global nitrogen cycle. Nitrogenase, the enzyme responsible for dinitrogen reduction, requires large amounts of ATP and reducing equivalents. Consequently, the nitrogen fixation process is subjected to sophisticated regulatory networks that respond to multiple environmental stimuli. In the free-living photosynthetic nitrogen-fixing bacterium Rhodospirillum rubrum, the activity of nitrogenase is tightly regulated at the post-translational level by reversible ADP-ribosylation in response to cellular changes in nitrogen and energy status, the so-called “switch-off” effect. Our studies have been focused on identifying the intracellular signal(s) and protein components acting during “switch-off”, and elucidating the mechanism underlying this regulation. We have shown that PII signal transduction proteins and the ammonium transporter AmtB1 play central roles in the signal transduction pathway leading to the post-translational regulation of nitrogenase, in particular, the involvement of AmtB1-PII complex formation during ammonium “switch-off”. In contrast, a different signaling pathway is operating during the energy “switch-off”, and several interesting differences are highlighted here. In addition, we have solved a high-resolution structure of Dinitrogenase Reductase Activating Glycohydrolase (DRAG) using X-ray crystallography. A detailed mechanism of ADP-ribose removal by DRAG is proposed, with our structural and functional studies on DRAG supporting a reversible membrane association mechanism of regulating its activity, further controlling the activity of nitrogenase.

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