Project description:EMG produced TPA metagenomics assembly of the Molecular study on haloarchaea producing industerially important enzymes (Molecular study on Haloarchaea) data set
Project description:We used quantitative proteomics (8-plex iTRAQ labeling) to investigate pathways and enzymes potentially important for sucrose metabolism in the halophilic archaeon Halohasta litchfieldiae. The data provided insights into the dynamics of the Hht. litchfieldiae proteome in response to sucrose, advancing the understanding of sucrose and fructose metabolism in haloarchaea.
Project description:In hypersaline brines, biodegradation of recalcitrant plant polymers can be inhibited by salt-induced microbial stress and/or caused by inadequate metabolic capabilities of extremely halophilic microbes. Therefore, woody materials can be well-preserved even in NaCl brines that are less biologically hostile than most other brines. Here, we considered whether the nanohaloarchaea, that live alongside (the related) haloarchaea, ever partake in the degradation of xylan, a major hemicellulose component of wood. Samples were taken from natural evaporitic brines and anthropogenic solar salterns located in various parts of Europe and Asia. We recently demonstrated that nanohaloarchaeon Ca. Nanohalobium constans lives as an ectosymbiont associated with the chitinolytic haloarchaeon Halomicrobium. Here, we describe an extremely halophilic xylan-degrading consortium with three members, where nanohaloarchaea act as ectosymbionts of Haloferax lucertensis, which in turn acts as a scavenger of xylan-degradation products, produced by a primary xylan hydrolytic Halorhabdus species. The two corresponding binary associations of nanohaloarchaea, Candidatus Nanohalococcus occultus SVXNc and Candidatus Nanohalovita haloferacivicina BNXNv and their hosts were obtained, stably cultivated and characterized. In contrast to the previously described association of chitinolytic haloarchaeon Halomicrobium and its amylolytic symbiont Ca. Nanohalobium, the host haloarchaea within the xylan-degrading consortium could metabolize α-glucans (glycogen and starch), and, thus, obtained no obvious trophic benefit from ectosymbionts. The current study has broadened the range of culturable ectosymbiontic nanohaloarchaea and demonstrates that they are an important ecophysiological component of polysaccharide-degrading halophilic microbial communities and can be readily isolated in binary co-cultures by using the appropriate enrichment strategy.
Project description:Small RNAs have been studied in detail in Bacteria and Eukarya domain, but in the case of Archaea domain the knowledge is scarce and the physiological function of the majority is still uncertain. To extend the knowledge of sRNAs in Archaea domain and its possible role in the regulation of the nitrogen assimilation metabolism in haloarchaea, Haloferax mediterranei has been used as a model microorganism. Bioinformatic approach has allowed to predict 295 putative sRNAs genes in the genome of H. mediterranei, 88 of which have been verified by means of RNA-seq. The secondary structure of putative sRNAs and its possible targets have been identified. Curiously, some of them present as possible targets genes related to the nitrogen assimilation, as glutamate dehydrogenase or regulatory nitrogen protein PII. Analysis of RNA-seq data has also revealed differences in the expression pattern of 16 sRNAs according to the nitrogen source. Consequently, RNomic and the bioinformatic approaches used in this work have allowed the identification of new sRNAs in Hfx. mediterranei, some of which show different expression pattern depending on the nitrogen source. It suggests that these sRNAs could be involved in the regulation of nitrogen assimilation, being able to constitute important gene regulatory network.
Project description:Oxidative stress responsive small non-coding RNAs (sRNAs) have been reported in the model archaeon, Haloferax volcanii, but targets and mechanisms of actions have not been elucidated. While haloarchaea are highly resistant to oxidative stress, a comprehensive understanding of the mechanisms regulating this remarkable response is lacking. Here, using a combination of high throughput and reverse molecular genetic approaches we elucidated the functional role of the most up-regulated intergenic sRNA during oxidative stress in H. volcanii, aptly named Small RNA in Haloferax Oxidative Stress (SHOxi). We demonstrated that SHOxi is a functional non-coding RNA that plays gene regulatory roles in the oxidative stress response of an extremophilic archaeon. We found that SHOxi likely regulates redox homeostasis during oxidative stress by the post-transcriptional destabilization of malic enzyme mRNA. The decrease in the NAD+/NADH ratio resulting from the direct RNA-RNA interaction between SHOxi and its trans-target is instrumental in the survival of H. volcanii. The regulatory effects of SHOxi provides evidence that the fine-tuning of metabolic cofactors could be a core strategy to mitigate damage from oxidative stress and confer resistance. This study is the first to establish the regulatory effects of sRNAs on mRNAs during the oxidative stress response in Archaea.
Project description:Dimitry Y. Sorokin et al., (2021, Russian Academy of Sciences, Russia and Delft University of Technology, The Netherlands) describe the isolation and physiological and genomic properties of a fifth functional group of sulfur-respiring haloarchaea enriched from hypersaline lake sediments with CO as the electron donor. Additional shotgun proteomic profiling of the described strains has been performed.
Project description:Many proteins and enzymes involved in denitrification in haloarchaea can be inferred to be located between the cytoplasmic membrane and the S-layer, based on the presence of a Tat signal sequence and the orientation of the active site that some of these enzymes have. The membrane fraction of the haloarchaeon Haloferax mediterranei (R-4), grown under anaerobic conditions in the presence of nitrate, was solubilised to identify the respiratory proteins associated or anchored to it. Using Triton X-100, CHAPS and n-Octyl-β-d-glucopyranoside at different concentrations we found the best conditions for isolating membrane proteins in micelles, in which enzymatic activity and stability were maintained. Then, they were subjected to purification using two chromatographic steps followed by the analysis of the eluents by NANO-ESI Chip-HPLC-MS/MS. The results showed that the four main enzymes of denitrification (nitrate, nitrite, nitric oxide and nitrous oxide reductases) in H. mediterranei were identified and they were co-purified thanks to the micelles made with Triton X-100 (20% w/v for membrane solubilisation and 0.2% w/v in the buffers used during purification). In addition, several accessory proteins involved in electron transfer processes during anaerobic respiration as well as proteins supporting ATP synthesis, redox balancing and oxygen sensing were detected. This is the first characterization of anaerobic membrane proteome of haloarchaea under denitrifying conditions using liquid chromatography-mass spectrometry. It provides new information for a better understanding of the anaerobic respiration in haloarchaea.