NifQ and NifO are essential to express nitrogenase activity in the presence of nitrate in Azotobacter vinelandii

Jiménez Vicente, Emilio and Barahona Martín, Emma and Wilcoxen, J. and Navarro Rodríguez, Mónica and Buesa Galiano, José María and Britt, David and Rubio Herrero, Luis Manuel (2015). NifQ and NifO are essential to express nitrogenase activity in the presence of nitrate in Azotobacter vinelandii. In: "XV Congress of the Spanish Society of Nitrogen Fixation, and the IV Portuguese-Spanish Congress on Nitrogen Fixation", 16/06/2015-18/06/2015, León, España. p. 1.

Description

Title: NifQ and NifO are essential to express nitrogenase activity in the presence of nitrate in Azotobacter vinelandii
Author/s:
  • Jiménez Vicente, Emilio
  • Barahona Martín, Emma
  • Wilcoxen, J.
  • Navarro Rodríguez, Mónica
  • Buesa Galiano, José María
  • Britt, David
  • Rubio Herrero, Luis Manuel
Item Type: Presentation at Congress or Conference (Article)
Event Title: XV Congress of the Spanish Society of Nitrogen Fixation, and the IV Portuguese-Spanish Congress on Nitrogen Fixation
Event Dates: 16/06/2015-18/06/2015
Event Location: León, España
Title of Book: XV Congress of the Spanish Society of Nitrogen Fixation, and the IV Portuguese-Spanish Congress on Nitrogen Fixation
Date: 2015
Subjects:
Faculty: Centro de Investigación en Biotecnología y Genómica de Plantas (CBGP) (UPM)
Department: Biotecnología - Biología Vegetal
Creative Commons Licenses: Recognition - No derivative works - Non commercial

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Abstract

In the presence of nitrate, Azotobacter vinelandii is able to assimilate nitrogen by using nitrogenase and nitrate reductase/nitrite reductase pathways simultaneously. Nitrogenase and nitrate reductase are Mo-enzymes containing FeMo-co and Mo-MGD at their active sites, respectively. In order to optimize the use of Mo, a scarce metal in nature, regulation of Mo distribution between both enzymes must be strictly controlled during nitrogen assimilation processes. The nifO and nifQ genes are grouped together with nifB, fdxN and rhdN in one transcriptional unit. It has been shown that nifO and nifQ expression levels change antagonistically depending on the presence of Mo in the medium (Rodriguez-Quinones 1993). In addition, the nifO mutant exhibits a Nif- phenotype in the presence of nitrate, whereas nifO overexpression lowers nitrate reductase activity (Gutierrez, J.C. 1997). The nifQ mutant is unable to fix N2 unless growth medium is supplemented with 1000-fold excess of Mo. Importantly, NifQ has been characterized as the physiological Mo donor to a NifEN/NifH complex during FeMo-co synthesis. (Hernandez, J.A. 2008). We aimed to understand the relationship between NifO and NifQ during expression of nitrogenase activity in presence of nitrate in A. vinelandii. The nifQ mutant was unable to fix N2 in the presence of nitrate, independently of the level of Mo in the medium. In contrast nifQ mutant showed enhanced nitrate reductase activity. Analysis of nitrogenase and nitrate reductase activities demonstrated that the nifQ overexpressing strain exhibited lower nitrogenase activity and higher nitrate reductase activity than wild-type when grown diazotrophically in the presence of nitrate, a phenotype similar to the nifO mutant (Gutierrez, J.C. 1997). An antagonist effect had been observed in the nifO overexpressing strain (Gutierrez, J.C. 1997). Simultaneous overexpression of both nifQ and nifO yielded nitrogenase and nitrate reductase activities similar to wild-type. The phenotype observed in nifQ overexpressing, but not in nifOQ overexpressing strain, points to NifO as candidate to preserve NifQ as Mo donor to nitrogenase when nitrate reductase is present. Transcriptional expression analysis performed by RT-qPCR showed lower expression of nitrogenase structural genes in the nifO mutant. In contrast increased expression of nitrate and nitrite reductase structural genes was observed for both nifO mutant and nifQ overexpression strains. Comparison between NifQ proteins isolated before and after addition of nitrate to the same culture of a nifQ overexpressing strain grown under diazotrophic conditions, showed NifQ cluster content alteration, resulting in decrease of [Mo-3Fe-4S]3+ and increase of [3Fe-4S]+ clusters. This effect of nitrate is consistent with the inability of NifQ to donate Mo for FeMo-co biosynthesis under nitrate reductase derepressing conditions. These results revealed two Mo pathways to nitrogenase: one that can be sorted by a large excess of Mo in the medium, and a second pathway strictly dependent on NifQ and NifO that would be essential to maintain active nitrogenase while assimilating nitrate through the molybdoenzyme nitrate reductase.

More information

Item ID: 42297
DC Identifier: http://oa.upm.es/42297/
OAI Identifier: oai:oa.upm.es:42297
Official URL: http://congreso.xvsefin.unileon.es/
Deposited by: Memoria Investigacion
Deposited on: 27 Oct 2016 13:33
Last Modified: 27 Oct 2016 13:33
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