Archaeal tRNA-Splicing Endonuclease as an Effector for RNA Recombination and Novel Trans-Splicing Pathways in Eukaryotes

We have characterized a homodimeric tRNA endonuclease from the euryarchaeota Ferroplasma acidarmanus (FERAC), a facultative anaerobe which can grow at temperatures ranging from 35 to 42 °C. This enzyme, contrary to the eukaryal tRNA endonucleases and the homotetrameric Methanocaldococcus jannaschii (METJA) homologs, is able to cleave minimal BHB (bulge–helix–bulge) substrates at 30 °C. The expression of this enzyme in Schizosaccharomyces pombe (SCHPO) enables the use of its properties as effectors by inserting BHB motif introns into hairpin loops normally seen in mRNA transcripts. In addition, the FERAC endonuclease can create proteins with new functionalities through the recombination of protein domains.

Microbial Population of Industrial Biooxidation Reactors

Microbial population performing biooxidation of flotation concentrate of gold bearing sulfide ore containing pyrite, arsenopyrite, and pyrrhotite was studied using cultural and molecular biological (metagenomics sequencing of V3-V4 fragments of 16S rRNA gene) approaches. The biooxidation of the concentrate was conducted at temperatures from 38 to 42°C. Strains of Acidithiobacillus thiooxidans, Acidiphilium multivorum, Leptospirillum ferriphilum, Sulfobacillus thermotolerans, Ferroplasma acidarmanus, and Ferroplasma acidiphilum were isolated from the samples of the pulp from biooxidation reactors. It was shown that optimum temperatures of isolated strains were from 38 to 40°C. Metagenomic analysis demonstrated predominance of the genera Acidiferrobacter, Acidithiobacillus, Acidiphilum, Leptospirillum, and Ferroplasma. According to results of molecular biological analysis, share of the genus Acidithiobacillus was of 0 to 25%, share of the genus Acidiferrobacter was of 7 to 56%, share of the genus Acidiphilum was of 0.03 to 36%, share of the genus Leptospirillum was of 0.7 to 7%, whereas share of the archaea of the genus Ferroplasma was of 33 to 94%. Thus, it was shown that representatives of the genus Ferroplasma can play significant role in bioleach process. Representatives of the genus Acidiferrobacter were previously detected in acid mine drainages, acid soils as well as in bioleach heaps and reactors, whereas data on predominance of the genus in tank bioleach processes have not been presented in the literature. In the present study, strains of the genus Acidiferrobacter were not isolated despite application of the nutrient media recommended for Acidiferrobacter and their properties were not studied. Nevertheless, results of the present study suggest that representatives of the genus Acidiferrobacter have a great impact on industrial bioleach processes.

Molybdate treatment and sulfate starvation decrease ATP and DNA levels inFerroplasma acidarmanus

Sulfate is a primary source of sulfur for most microbes and in some prokaryotes it is used an electron acceptor. The acidophileFerroplasma acidarmanus(strain fer1) requires a minimum of 150 mM of a sulfate-containing salt for growth. Sulfate is assimilated byF. acidarmanusinto proteins and reduced to form the volatile organic sulfur compounds methanethiol and dimethyldisulfide. In the absence of sulfate, cell death occurs by an unknown mechanism. In this study, cell viability and genomic DNA and ATP contents ofF. acidarmanuswere monitored in response to the absence of sulfate or the presence of sulfate and the sulfate analog molybdate (MoO42-). Cellular DNA and ATP contents were monitored as markers of cell viability. The absence of sulfate led to a decrease in viable cell numbers of greater than 7 log10within 5 days, a > 99% reduction in genomic DNA within 3 days, and a > 60% decrease in ATP within 6 h. Likewise, cells incubated with lost viability (decreased by > 2 log10in 5 days), extractable genomic DNA (reduction of > 60% in 2 days), and ATP (reduction of > 70 % in 2 hours). These results demonstrate that sulfate deprivation or the presence of molybdate have similar impacts on cell viability and essential biomolecules. Sulfate was coupled to cellular ATP content and maintenance of DNA integrity inF. acidarmanus, a finding that may be applicable to other acidophiles that are typically found in sulfate-rich biotopes.