Extracellular membrane vesicles in the three domains of life and beyond.

Cells from all three domains of life, Archaea, Bacteria and Eukarya, produce extracellular vesicles (EVs) which are sometimes associated with filamentous structures known as nanopods or nanotubes. The mechanisms of EV biogenesis in the three domains remain poorly understood, although studies in Bacteria and Eukarya indicate that the regulation of lipid composition plays a major role in initiating membrane curvature. EVs are increasingly recognized as important mediators of intercellular communication via transfer of a wide variety of molecular cargoes.

Greigite nanocrystals produced by hyperthermophilic archaea of Thermococcales order.

Interactions between hyperthermophilic archaea and minerals occur in hydrothermal deep-sea vents, one of the most extreme environments for life on Earth. These interactions occur in the internal pores and at surfaces of active hydrothermal chimneys. In this study, we show that, at 85°C, Thermococcales, the predominant hyperthermophilic microorganisms inhabiting hot parts of hydrothermal deep-sea vents, produce greigite nanocrystals (Fe3S4) on extracellular polymeric substances, and that an amorphous iron phosphate acts as a precursor phase.

Synergistic template-free synthesis of dsDNA by Thermococcus nautili primase PolpTN2, DNA polymerase PolB, and pTN2 helicase.

A combination of three enzymes from the hyperthermophilic archaeon Thermococcus nautili, DNA primase PolpTN2, DNA polymerase PolB, and pTN2 DNA helicase, was found to synthesize up to 300-400 ng/µl dsDNA from deoxynucleotide triphosphates in less than 30 min in the absence of added template DNA and oligonucleotide primer. The reaction did not occur below 64 °C. No synthesis was observed if PolpTN2 or PolB were left out; helicase was not essential but accelerated the reaction.

The SF1 helicase encoded by the archaeal plasmid pTN2 of Thermococcus nautili.

We expressed, purified, and characterized the helicase encoded by ORF1 of the Thermococcus nautili pTN2 plasmid (Soler et al. Nucl Acids Res 38, 5088-5104, 2010). The enzyme, which belongs to the SF1 family of helicases, possesses NTPase activity, with a strong preference for ATP and GTP as compared to CTP and TTP; dATP was also a substrate.

A highly divergent archaeo-eukaryotic primase from the Thermococcus nautilus plasmid, pTN2.

We report the characterization of a DNA primase/polymerase protein (PolpTN2) encoded by the pTN2 plasmid from Thermococcus nautilus. Sequence analysis revealed that this protein corresponds to a fusion between an N-terminal domain homologous to the small catalytic subunit PriS of heterodimeric archaeal and eukaryotic primases (AEP) and a C-terminal domain related to their large regulatory subunit PriL. This unique domain configuration is not found in other virus- and plasmid-encoded primases in which PriS-like domains are typically fused to different types of helicases.

Interaction of the family-B DNA polymerase from the archaeon Pyrococcus furiosus with deaminated bases.

The interaction of archaeal family B DNA polymerases with deaminated bases has been examined. As determined previously by our group, the polymerase binds tightly to uracil (the deamination product of cytosine), in single-stranded DNA, and stalls replication on encountering this base. DNA polymerisation was also inhibited by the presence of hypoxanthine, the deamination product of adenine.