Abstract
The photosynthetic bacterium Rhodopseudomonas palustris converts nitrogen gas (N2) to fertilizer ammonia (NH3) and also produces clean energy hydrogen gas (H2) from protons (H+) when it is grown anaerobically in nitrogen fixing medium with illumination, a condition that promotes the expression of active nitrogenase. Compared with quantitative real-time PCR (qRT-PCR) and the lacZ reporter system, two methods commonly used for in vivo study of nitrogenase regulation in photosynthetic bacteria, the fluorescent protein reporter system has advantages in terms of its simplicity and sensitivity. However, little is known concerning if the fluorescent protein reporter system can be used in bacterial cells that need to grow anaerobically. Here, we developed an RFP-based method to measure the nitrogenase gene expression in photosynthetic bacteria grown anaerobically. This method was able to determine the levels of both the genome-based and the plasmid-based nitrogenase expression under anaerobic conditions, providing a better method for in vivo study of gene expression affected by oxygen. The RFP reporter system developed here will promote a better understanding of the molecular mechanism of nitrogenase regulation and will be used on other genes of interest in a wider range of anaerobic bacteria.
Highlights
Accepted: 12 January 2022Rhodopseudomonas palustris is a photosynthetic nitrogen-fixing bacterium widely distributed in nature [1]
R. palustris, one of the most important model organisms for in vivo studies of nitrogenases, generates a large amount of ATP required for nitrogen fixation from light energy under anaerobic conditions [1,32,33]
To examine if red fluorescent protein (RFP) still can fluoresce when it is switched from anaerobic to aerobic environments, R. palustris CGA3005 cells expressing RFP were taken from the sealed culture tubes and their fluorescence intensities were measured
Summary
Accepted: 12 January 2022Rhodopseudomonas palustris is a photosynthetic nitrogen-fixing bacterium widely distributed in nature [1]. Diazotrophs convert atmospheric nitrogen gas (N2 ) to ammonia (NH3 ) by nitrogenase as a major source of bioavailable nitrogen. V nitrogenase, and iron-iron (FeFe) or Fe-only nitrogenase, each differentiated by the metal composition in the active site cofactor [3,4]. In addition to nitrogen fixation and hydrogen production, Fe-only nitrogenase is capable of reducing carbon dioxide (CO2 ) to methane (CH4 ), bringing a much stronger greenhouse gas (CH4 ) to the atmosphere and causing an acceleration of global warming [5]. R. palustris is able to express all three nitrogenases, making it an attractive model bacterium for in vivo studies of nitrogenase [5,6]. R. palustris expresses nitrogenase and fixes nitrogen only when grown anaerobically with illumination ( it grows well aerobically in the medium supplemented with fixed nitrogen such as ammonia).
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call