Small-Molecule Mn Antioxidants in Caenorhabditis elegans and Deinococcus radiodurans Supplant MnSOD Enzymes during Aging and Irradiation

The current theory of cellular defense against oxidative damage identifies antioxidant enzymes as primary defenders against ROS, with MnSOD being the preeminent superoxide (O 2 •− ) scavenger. However, MnSOD is shown to be dispensable both for radiation resistance and longevity in model organisms, the bacterium Deinococcus radiodurans and the nematode Caenorhabditis elegans .

Glycan profiling of the gut microbiota by Glycan-seq

Bacterial glycans modulate the cross talk between the gut microbiota and its host. However, little is known about these glycans because of the lack of appropriate technology to study them. In this study, we applied Glycan-seq technology for glycan profiling of the intact gut microbiota of mice. The evaluation of cultured gram-positive (Deinococcus radiodurans) and gram-negative (Escherichia coli) bacteria showed significantly distinct glycan profiles between these bacteria, which were selected and further analyzed by flow cytometry. The results of flow cytometry agreed well with those obtained by Glycan-seq, indicating that Glycan-seq can be used for bacterial glycan profiling. We thus applied Glycan-seq for comparative glycan profiling of pups and adult mice gut microbiotas. The glycans of the pups and adult microbiotas had significantly distinct glycan profiles, which reflect the different bacterial compositions of pups and adult gut microbiotas based on 16S rRNA gene sequencing.α2-6Sia-binders bound specifically to the pups microbiota. Lectin pull-down followed by 16S rRNA gene sequencing of the pups microbiota identified Lactobacillaceae as the most abundant bacterial family with glycans reacting with α2-6Sia-binders. The Glycan-seq system can reveal the glycan profile of the intact bacterial gut microbiota.

Proteomic Response of Deinococcus radiodurans to Short-Term Real Microgravity during Parabolic Flight Reveals Altered Abundance of Proteins Involved in Stress Response and Cell Envelope Functions

Rapidly evolving space exploration makes understanding the short- and long- term effects of microgravity on humans, plants, and microorganisms an important task. The ubiquitous presence of the gravitational force has had an influence on the development of all living entities on Earth, and short- and long-term changes in perceived gravitational force can induce notable changes within cells. Deinococcus radiodurans is the Gram-positive bacterium that is best known for its extreme resistance to UV-C and gamma radiation, oxidation stress, and desiccation. Thus increased interest has been placed on this species in the context of space research. The present study aims to elucidate the short-term proteomic response of this species to real microgravity during parabolic flight. Overnight cultures of D. radiodurans were subjected to microgravity during a single parabola, and metabolic activity was quenched using methanol. Proteins were extracted and subsequently measured using HPLC nESI MS/MS. The results, such as the enrichment of the peptidoglycan biosynthesis pathway with differentially abundant proteins and altered S-layer protein abundance, suggested molecular rearrangements in the cell envelope of D. radiodurans. Altered abundance of proteins involved in energy metabolism and DNA repair could be linked with increased endogenous ROS production that contributes to the stress response. Moreover, changes in protein abundance in response to microgravity show similarities with previously reported stress responses. Thus, the present results could be used to further investigate the complex regulation of the remarkable stress management of this bacterium.

Deinococcus radiodurans Exopolysaccharide Inhibits Staphylococcus aureus Biofilm Formation

Deinococcus radiodurans is an extremely resistant bacterium against extracellular stress owing to on its unique physiological functions and the structure of its cellular constituents. Interestingly, it has been reported that the pattern of alteration in Deinococcus proportion on the skin is negatively correlated with skin inflammatory diseases, whereas the proportion of Staphylococcus aureus was increased in patients with chronic skin inflammatory diseases. However, the biological mechanisms of deinococcal interactions with other skin commensal bacteria have not been studied. In this study, we hypothesized that deinococcal cellular constituents play a pivotal role in preventing S. aureus colonization by inhibiting biofilm formation. To prove this, we first isolated cellular constituents, such as exopolysaccharide (DeinoPol), cell wall (DeinoWall), and cell membrane (DeinoMem), from D. radiodurans and investigated their inhibitory effects on S. aureus colonization and biofilm formation in vitro and in vivo. Among them, only DeinoPol exhibited an anti-biofilm effect without affecting bacterial growth and inhibiting staphylococcal colonization and inflammation in a mouse skin infection model. Moreover, the inhibitory effect was impaired in the Δdra0033 strain, a mutant that cannot produce DeinoPol. Remarkably, DeinoPol not only interfered with S. aureus biofilm formation at early and late stages but also disrupted a preexisting biofilm by inhibiting the production of poly-N-acetylglucosamine (PNAG), a key molecule required for S. aureus biofilm formation. Taken together, the present study suggests that DeinoPol is a key molecule in the negative regulation of S. aureus biofilm formation by D. radiodurans. Therefore, DeinoPol could be applied to prevent and/or treat infections or inflammatory diseases associated with S. aureus biofilms.

Probing the molecular mechanism of action of Deinococcus radiodurans RecD2 at single-molecule resolution

Helicases are ATP-driven molecular machines that directionally remodel nucleic acid polymers in all three domains of life. Helicases are responsible for resolving double-stranded DNA (dsDNA) into separate single-strands and this activity is essential for DNA replication, nucleotide excision repair, and homologous recombination. RecD2 from Deinococcus radiodurans (DrRecD2) has important contributions towards its unusually high tolerance to gamma radiation and hydrogen peroxide. Although previous X-ray Crystallography studies have revealed the structural characteristics of the protein, the direct experimental evidence regarding the dynamics of the DNA unwinding process by DrRecD2 in the context of other accessory proteins is yet to be found. In this study, we have probed the exact binding event and processivity of DrRecD2 at single-molecule resolution using Protein-induced fluorescence enhancement (smPIFE) and Forster resonance energy transfer (smFRET). We have found that the protein prefers to bind at the 5 prime terminal end of the single-stranded DNA (ssDNA) by Drift and has helicase activity even in absence of ATP. However, a faster and iterative mode of DNA unwinding was evident in presence of ATP. The rate of translocation of the protein was found to be slower on dsDNA compared to ssDNA. We also showed that DrRecD2 is recruited at the binding site by the single-strand binding protein (SSB) and during the unwinding, it can displace RecA from ssDNA.

Enhancing the Inhomogeneonus Photodynamics of Canonical Bacteriophytochrome

The ability of phytochromes to act as photoswitches in plants and microorganisms depends on interactions between a bilin-like chromophore and a protein. The interconversion occurs between the spectrally distinct red (Pr) and far-red (Pfr) conformers. This conformational change is triggered by the photoisomerization of the chromophore D-ring pyrrole. In this study, as a representative example of a phytochrome-bilin system, we take biliverdin IXα (BV) bound to bacteriophytochrome (BphP) from Deinococcus radiodurans. In the absence of light, we use an enhanced sampling molecular dynamics (MD) method to overcome the photoisomerization energy barrier. We find that the calculated free energy (FE) barriers between essential metastable states agree with spectroscopic results. We show that the enhanced dynamics of the BV chromophore in BphP triggers nanometer-scale conformational movements that propagate by two experimentally determined signal transduction pathways. Most importantly, we describe how the metastable states enable a thermal transition known as the dark reversion between Pfr and Pr, through a previously unknown intermediate state of Pfr. Here, for the first time, the heterogeneity of temperature-dependent Pfr states is presented at the atomistic level. This work paves a way toward understanding the complete mechanism of the photoisomerization of a bilin-like chromophore in phytochromes.

The journey of Deinococcus radiodurans; a perspective

: Deinococcus radiodurans has been recognized for its robustness and recorded in the Guinness Book of World Records as the world's toughest known bacterium. In essence, the title comes from its ability to survive extreme conditions such as severe drought (desiccation) and radiation tolerance up to 15000 Gy, which is more than 250 times of E. coli and about 3000 times of humans. Due to its high tolerance to all kinds of genotoxic stress, such as desiccation, UV, X-rays, and oxidants, D. radiodurans is a well-suited model organism for microbial radiation resistance studies. The DNA damage-responsive gene expression is an important component of post-stress recovery where the cell shows a great multiplicity of genomes leading to the highly proficient recombinational DNA repair. This article pitches light on the unique properties of D. radiodurans, unfolding its journey so far as well as important molecular discoveries, prospects, and biotechnological applications.

Enhancing the Inhomogeneonus Photodynamics of Canonical Bacteriophytochrome

The ability of phytochromes to act as photoswitches in plants and microorganisms depends on interactions between a bilin-like chromophore and a protein. The interconversion occurs between the spectrally distinct red (Pr) and far-red (Pfr) conformers. This conformational change is triggered by the photoisomerization of the chromophore D-ring pyrrole. In this study, as a representative example of a phytochrome-bilin system, we take biliverdin IXα (BV) bound to bacteriophytochrome (BphP) from Deinococcus radiodurans. In the absence of light, we use an enhanced sampling molecular dynamics (MD) method to overcome the photoisomerization energy barrier. We find that the calculated free energy (FE) barriers between essential metastable states agree with spectroscopic results. We show that the enhanced dynamics of the BV chromophore in BphP triggers nanometer-scale conformational movements that propagate by two experimentally determined signal transduction pathways. Most importantly, we describe how the metastable states enable a thermal transition known as the dark reversion between Pfr and Pr, through a previously unknown intermediate state of Pfr. Here, for the first time, the heterogeneity of temperature-dependent Pfr states is presented at the atomistic level. This work paves a way toward understanding the complete mechanism of the photoisomerization of a bilin-like chromophore in phytochromes.

DNA Damage Protection for Enhanced Bacterial Survival Under Simulated Low Earth Orbit Environmental Conditions in Escherichia coli

Some organisms have shown the ability to naturally survive in extreme environments, even outer space. Some of these have natural mechanisms to resist severe DNA damage from conditions such as ionizing and non-ionizing radiation, extreme temperatures, and low pressures or vacuum. A good example can be found in Deinococcus radiodurans, which was exposed to severe conditions such as those listed in the Exposure Facility of the International Space Station (ISS) for up to three years. Another example are tardigrades (Ramazzottius varieornatus) which are some of the most resilient animals known. In this study, the survival under simulated Low earth Orbit (LEO) environmental conditions was tested in Escherichia coli. The radiation resistance of this bacteria was enhanced using the Dsup gene from R. varieornatus, and two more genes from D. radiodurans involved in DNA damage repair, RecA and uvrD. The enhanced survival to wide ranges of temperatures and low pressures was then tested in the new strains. This research constitutes a first step in the creation of new bacterial strains engineered to survive severe conditions and adapting existing species for their survival in remote environments, including extra-terrestrial habitats. These strains could be key for the development of environments hospitable to life and could be of use for ecological restoration and space exploration. In addition, studying the efficacy and the functioning of the DNA repair mechanisms used in this study could be beneficial for medical and life sciences engineering.