Diazotrophic Growth Allows Azotobacter vinelandii To Overcome the Deleterious Effects of a glnE Deletion release_yanwquzvfbhm5e5sietnnfmgbu

by Florence Mus, Alex Tseng, Ray Dixon, John W. Peters

Published in Applied and Environmental Microbiology by American Society for Microbiology.

2017   Volume 83, Issue 13

Abstract

<jats:title>ABSTRACT</jats:title> Overcoming the inhibitory effects of excess environmental ammonium on nitrogenase synthesis or activity and preventing ammonium assimilation have been considered strategies to increase the amount of fixed nitrogen transferred from bacterial to plant partners in associative or symbiotic plant-diazotroph relationships. The GlnE adenylyltransferase/adenylyl-removing enzyme catalyzes reversible adenylylation of glutamine synthetase (GS), thereby affecting the posttranslational regulation of ammonium assimilation that is critical for the appropriate coordination of carbon and nitrogen assimilation. Since GS is key to the sole ammonium assimilation pathway of <jats:named-content content-type="genus-species">Azotobacter vinelandii</jats:named-content> , attempts to obtain deletion mutants in the gene encoding GS ( <jats:italic>glnA</jats:italic> ) have been unsuccessful. We have generated a <jats:italic>glnE</jats:italic> deletion strain, thus preventing posttranslational regulation of GS. The resultant strain containing constitutively active GS is unable to grow well on ammonium-containing medium, as previously observed in other organisms, and can be cultured only at low ammonium concentrations. This phenotype is caused by the lack of downregulation of GS activity, resulting in high intracellular glutamine levels and severe perturbation of the ratio of glutamine to 2-oxoglutarate under excess-nitrogen conditions. Interestingly, the mutant can grow diazotrophically at rates comparable to those of the wild type. This observation suggests that the control of nitrogen fixation-specific gene expression at the transcriptional level in response to 2-oxoglutarate via NifA is sufficiently tight to alone regulate ammonium production at levels appropriate for optimal carbon and nitrogen balance. <jats:bold>IMPORTANCE</jats:bold> In this study, the characterization of the <jats:italic>glnE</jats:italic> knockout mutant of the model diazotroph <jats:named-content content-type="genus-species">Azotobacter vinelandii</jats:named-content> provides significant insights into the integration of the regulatory mechanisms of ammonium production and ammonium assimilation during nitrogen fixation. The work reveals the profound fidelity of nitrogen fixation regulation in providing ammonium sufficient for maximal growth but constraining energetically costly excess production. A detailed fundamental understanding of the interplay between the regulation of ammonium production and assimilation is of paramount importance in exploiting existing and potentially engineering new plant-diazotroph relationships for improved agriculture.
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Type  article-journal
Stage   published
Date   2017-04-21
Language   en ?
DOI  10.1128/aem.00808-17
PubMed  28432097
PMC  PMC5478974
Wikidata  Q49215815
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