Analysis of Haloferax mediterranei Lrp Transcriptional Regulator

• Published in: Genes 2021-05, Vol.12 (6), p.802
• Main Authors: Matarredona, Laura, Camacho, Mónica, García-Bonete, María-José, Esquerra, Belén, Zafrilla, Basilio, Esclapez, Julia, Bonete, María-José
• Format: Article
• Language: English
• Subjects: Amino acids; Amino Acids - metabolism; Ammonia - metabolism; Ammonium; Archaea; Archaeal Proteins - genetics; Archaeal Proteins - metabolism; Bacteria; Biological Sciences; Biologiska vetenskaper; Biotechnology; Carbon; Chromatography; Cloning; E coli; Enzymes; Gene expression; Gene Expression Regulation, Archaeal; Gene regulation; Genome, Archaeal; Genomes; Glutamate-ammonia ligase; Glutamine; Growth conditions; Haloferax mediterranei; Haloferax mediterranei - genetics; Haloferax mediterranei - metabolism; Heavy metals; His-tag; Homologous overexpression; Hydrogen peroxide; Ligands; Lrp; LRP protein; Metabolism; Metabolites; Microorganisms; Molecular modelling; Nickel; Nitrogen; Nitrogen - metabolism; Nitrogen cycle; Nitrogen sources; Phylogenetics; Plasmids; Promoter Regions, Genetic; Proteins; Reporter gene; Seawater; Starvation; Stress; Stress, Physiological; Transcription Factors - genetics; Transcription Factors - metabolism; Western blot; Western blotting; β-galactosidase assay
• ISSN: 2073-4425; 2073-4425
• DOI: 10.3390/genes12060802

is an extremely halophilic archaeon, able to live in hypersaline environments with versatile nutritional requirements, whose study represents an excellent basis in the field of biotechnology. The transcriptional machinery in combines the eukaryotic basal apparatus and the bacterial regulation mechanisms. However, little is known about molecular mechanisms of gene expression regulation compared with , particularly in Haloarchaea. The genome of contains a gene, (HFX_RS01210), which encodes a transcriptional factor belonging to Lrp/AsnC family. It is located downstream of the glutamine synthetase gene (HFX_RS01205), an enzyme involved in ammonium assimilation and amino acid metabolism. To study this transcriptional factor more deeply, the gene has been homologously overexpressed and purified under native conditions by two chromatographic steps, namely nickel affinity and gel filtration chromatography, showing that Lrp behaves asa tetrameric protein of approximately 67 kDa. Its promoter region has been characterized under different growth conditions using as a reporter gene. The amount of Lrp protein was also analyzed by Western blotting in different nitrogen sources and under various stress conditions. To sum up, regarding its involvement in the nitrogen cycle, it has been shown that its expression profile does not change in response to the nitrogen sources tested. Differences in its expression pattern have been observed under different stress conditions, such as in the presence of hydrogen peroxide or heavy metals. According to these results, the Lrp seems to be involved in a general response against stress factors, acting as a first-line transcriptional regulator.