Rhodamine6G and Hœchst33342 narrow BmrA conformational spectrum for a more efficient use of ATP.

Multidrug ABC transporters harness the energy of ATP binding and hydrolysis to translocate substrates out of the cell and detoxify them. While this involves a well-accepted alternating access mechanism, molecular details of this interplay are still elusive. Rhodamine6G binding on a catalytic inactive mutant of the homodimeric multidrug ABC transporter BmrA triggers a cooperative binding of ATP on the two identical nucleotide-binding-sites, otherwise michaelian.

Viral dynamics in a high-rate algal pond reveals a burst of diversity correlated with episodic algal mortality.

Unlabelled: This study explores virus-host dynamics in a unique environment: an industrial high-rate algal pond (HRAP). A wealth of novel DNA algal viruses are revealed, including members of "giant viruses" and the enigmatic (e.g.

The transport activity of the multidrug ABC transporter BmrA does not require a wide separation of the nucleotide-binding domains.

ATP-binding cassette (ABC) transporters are ubiquitous membrane proteins responsible for the translocation of a wide diversity of substrates across biological membranes. Some of them confer multidrug or antimicrobial resistance to cancer cells and pathogenic microorganisms, respectively. Despite a wealth of structural data gained in the last two decades, the molecular mechanism of these multidrug efflux pumps remains elusive, including the extent of separation between the two nucleotide-binding domains (NBDs) during the transport cycle.

CryoEM Data Analysis of Membrane Proteins. Practical Considerations on Amphipathic Belts, Ligands, and Variability Analysis.

Membrane proteins data analysis by cryoEM shows some specificities, as can be found in other typical investigations such as biochemistry, biophysics, or X-ray crystallography. Membrane proteins are typically surrounded by an amphipathic belt that will have some degree of influence on the 3D reconstruction and analysis. In this chapter, we review our experience with the ABC transporter BmrA, as well as our statistical analysis of amphipathic belts around membrane proteins, to bring awareness on some particular features of membrane protein investigations by cryoEM.

Evaluation of sequential filtration and centrifugation to capture environmental DNA and survey microbial eukaryotic communities in aquatic environments.

Sequential membrane filtration of water samples is commonly used to monitor the diversity of aquatic microbial eukaryotes. This capture method is efficient to focus on specific taxonomic groups within a size fraction, but it is time-consuming. Centrifugation, often used to collect microorganisms from pure culture, could be seen as an alternative to capture microbial eukaryotic communities from environmental samples.

Conformational space exploration of cryo-EM structures by variability refinement.

Cryo-EM observation of biological samples enables visualization of sample heterogeneity, in the form of discrete states that are separable, or continuous heterogeneity as a result of local protein motion before flash freezing. Variability analysis of this continuous heterogeneity describes the variance between a particle stack and a volume, and results in a map series describing the various steps undertaken by the sample in the particle stack. While this observation is absolutely stunning, it is very hard to pinpoint structural details to elements of the maps.

The non-Newtonian behavior of detergents during concentration is increased by macromolecules, in trans, and results in their over-concentration.

Concentration of pure membrane proteins in detergent solution results in detergent concentration, albeit in unknown amounts. This phenomenon is observed in every lab working on membrane proteins, but has seldom been investigated. In this study, we explored the behavior of detergents mixed with membrane proteins during the step of sample concentration using centrifugal devices.

Ecosystem-specific microbiota and microbiome databases in the era of big data.

The rapid development of sequencing methods over the past decades has accelerated both the potential scope and depth of microbiota and microbiome studies. Recent developments in the field have been marked by an expansion away from purely categorical studies towards a greater investigation of community functionality. As in-depth genomic and environmental coverage is often distributed unequally across major taxa and ecosystems, it can be difficult to identify or substantiate relationships within microbial communities.

Overexpression of the ABC Transporter BmrA Within Intracellular Caveolae in Escherichia coli.

We describe here the overproduction and oriented membrane insertion of membrane protein inside intracellular vesicles named heterologous caveolae within E. coli. The method is described with BmrA, a multidrug efflux pump from Bacillus subtilis.

Substrate-bound and substrate-free outward-facing structures of a multidrug ABC exporter.

Multidrug ABC transporters translocate drugs across membranes by a mechanism for which the molecular features of drug release are so far unknown. Here, we resolved three ATP-Mg-bound outward-facing conformations of the (homodimeric) BmrA by x-ray crystallography and single-particle cryo-electron microscopy (EM) in detergent solution, one of them with rhodamine 6G (R6G), a substrate exported by BmrA when overexpressed in . Two R6G molecules bind to the drug-binding cavity at the level of the outer leaflet, between transmembrane (TM) helices 1-2 of one monomer and TM5'-6' of the other.

A High Rate Algal Pond Hosting a Dynamic Community of RNA Viruses.

Despite a surge of RNA virome sequencing in recent years, there are still many RNA viruses to uncover-as indicated by the relevance of viral dark matter to RNA virome studies (i.e., putative viruses that do not match to taxonomically identified viruses).

CryoEM reconstructions of membrane proteins solved in several amphipathic solvents, nanodisc, amphipol and detergents, yield amphipathic belts of similar sizes corresponding to a common ordered solvent layer.

To maintain membrane proteins soluble in aqueous solution, amphipathic compounds are used to shield the hydrophobic patch of their membrane insertion, which forms a belt around the protein. This amphipathic belt is seldom looked at due to the difficulty to visualize it. Cryo-EM is now offering this possibility, where belts are visible in 3D reconstructions.

Accumulation of detached kelp biomass in a subtidal temperate coastal ecosystem induces succession of epiphytic and sediment bacterial communities.

Kelps are dominant primary producers in temperate coastal ecosystems. Large amounts of kelp biomass can be exported to the seafloor during the algal growth cycle or following storms, creating new ecological niches for the associated microbiota. Here, we investigated the bacterial community associated with the kelp Laminaria hyperborea during its accumulation and degradation on the seafloor.

Alterocin, an Antibiofilm Protein Secreted by sp. Strain 3J6.

We sought to identify and study the antibiofilm protein secreted by the marine bacterium sp. strain 3J6. The latter is active against marine and terrestrial bacteria, including clinical strains forming different biofilm types.

Degradation dynamics and processes associated with the accumulation of Laminaria hyperborea (Phaeophyceae) kelp fragments: an in situ experimental approach.

A high proportion of the kelp Laminaria hyperborea production is exported from kelp forests following seasonal storms or natural annual old blade loss. Transport of drifting kelp fragments can lead to temporary accumulations in benthic subtidal habitats. We investigated the degradation processes of L.

Evaluation of a new primer combination to minimize plastid contamination in 16S rDNA metabarcoding analyses of alga-associated bacterial communities.

Plant- and alga-associated bacterial communities are generally described via 16S rDNA metabarcoding using universal primers. As plastid genomes encode 16S rDNA related to cyanobacteria, these data sets frequently contain >90% plastidial sequences, and the bacterial diversity may be under-sampled. To overcome this limitation we evaluated in silico the taxonomic coverage for four primer combinations targeting the 16S rDNA V3-V4 region.

Evolutionary Evidence of Algal Polysaccharide Degradation Acquisition by 9 to Adapt to Macroalgal Niches.

About half of seaweed biomass is composed of polysaccharides. Most of these complex polymers have a marked polyanionic character. For instance, the red algal cell wall is mainly composed of sulfated galactans, agars and carrageenans, while brown algae contain alginate and fucose-containing sulfated polysaccharides (FCSP) as cell wall polysaccharides.

[Partial nephrectomy for renal masses >7cm: Morbidity, oncological and functional outcomes (UroCCR-7 study)].

Objectives: To describe the morbidity, mortality, oncological and functional results of Partial nephrectomy (PN) for the treatment of renal tumors of more than 7cm.

Material And Methods: Thirty-seven partial nephrectomies for tumors larger than 7cm operated in a single center between 1987 and 2016 were analyzed retrospectively. The pre, per and postoperative clinico-biological data were collected within the UroCCR database.

Seasonal and algal diet-driven patterns of the digestive microbiota of the European abalone Haliotis tuberculata, a generalist marine herbivore.

Background: Holobionts have a digestive microbiota with catabolic abilities allowing the degradation of complex dietary compounds for the host. In terrestrial herbivores, the digestive microbiota is known to degrade complex polysaccharides from land plants while in marine herbivores, the digestive microbiota is poorly characterized. Most of the latter are generalists and consume red, green, and brown macroalgae, three distinct lineages characterized by a specific composition in complex polysaccharides, which represent half of their biomass.

No difference in outcome for open versus arthroscopic rotator cuff repair: a prospective comparative trial.

Purpose: Arthroscopic techniques tend to become the gold standard in rotator cuff repair. However, little data are reported in the literature regarding the improvement of postoperative outcomes and re-tear rate relative to conventional open surgery. The aim of this study was to compare clinical outcomes and cuff integrity after arthroscopic versus open cuff repair.

The Complete Genome Sequence of the Fish Pathogen Provides Insights into Virulence Mechanisms.

is a devastating bacterial pathogen of wild and farmed marine fish with a broad host range and a worldwide distribution. We report here the complete genome sequence of the type strain NCIMB 2154. The genome consists of a 3,435,971-base pair circular chromosome with 2,866 predicted protein-coding genes.

Where less may be more: how the rare biosphere pulls ecosystems strings.

Rare species are increasingly recognized as crucial, yet vulnerable components of Earth's ecosystems. This is also true for microbial communities, which are typically composed of a high number of relatively rare species. Recent studies have demonstrated that rare species can have an over-proportional role in biogeochemical cycles and may be a hidden driver of microbiome function.

Polysaccharide utilisation loci of Bacteroidetes from two contrasting open ocean sites in the North Atlantic.

Marine Bacteroidetes have pronounced capabilities of degrading high molecular weight organic matter such as proteins and polysaccharides. Previously we reported on 76 Bacteroidetes-affiliated fosmids from the North Atlantic Ocean's boreal polar and oligotrophic subtropical provinces. Here, we report on the analysis of further 174 fosmids from the same libraries.