Is there a rate-limiting step in the catalytic cycle of [Ni-Fe] hydrogenases?

The question of the existence of a rate-limiting step in the catalytic cycle of Ni-Fe hydrogenases was taken up by using the sets of data available in the case of two specific enzymes: the hydrogenase from Thiocapsa roseopercisina, in which isotope effects have been systematically investigated over a wide pH range, and the enzyme from Desulfovibrio fructosovorans, for which the activities and the redox properties have been studied in two different forms, the wild type and the P238C mutant. When these data are analyzed in the light of appropriate kinetic models, it is concluded that electron transfer and proton transfer are rate limiting in the H2 uptake and H2 evolution reactions, respectively. This proposal is consistent with the data available from other Ni-Fe enzymes.     

Remote opto‐acoustic probing of single‐cell adhesion on metallic surfaces

The reflection of picosecond ultrasonic pulses from a cell‐substrate interface is used to probe cell‐biomaterial adhesion with a subcell resolution. We culture monocytes on top of a thin biocompatible Ti metal film, supported by a transparent sapphire substrate. Low‐energy femtosecond pump laser pulses are focused at the bottom of the Ti film to a micron spot. The subsequent ultrafast thermal expansion launches a longitudinal acoustic pulse in Ti, with a broad spectrum extending up to 100 GHz. We measure the acoustic echoes reflected from the Ti‐cell interface through the transient optical reflectance changes. The time‐frequency analysis of the reflected acoustic pulses gives access to a map of the cell acoustic impedance Z_c and to a map of the film‐cell interfacial stiffness K simultaneously. Variations in Z_c across the cell are attributed to rigidity and density fluctuations within the cell, whereas variations in K are related to interfacial intermolecular forces and to the nano‐architecture of the transmembrane bonds. (© 2014 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Fermentation procedure of a crude oil in continuous culture on seawater

Summary The crude oil was degraded (80%) in continuous culturing and non sterile conditions by a mixed bacteria community. The fermentation process relies on a step of pre-emulsification of the substrate before it is introduced into the reactor. The emulsification indispensable for degrading crude oil is performed by the mixed bacteria community during its growth on hydrocarbons. On the other hand, the use of an ultrafiltration device allows the obtention of high cell concentrations (7.6 g·l^-1) and high degradation rates.     

Detecting Signatures of Selection Through Haplotype Differentiation Among Hierarchically Structured Populations

The detection of molecular signatures of selection is one of the major concerns of modern population genetics. A widely used strategy in this context is to compare samples from several populations and to look for genomic regions with outstanding genetic differentiation between these populations. Genetic differentiation is generally based on allele frequency differences between populations, which are measured by F_ST or related statistics. Here we introduce a new statistic, denoted hapFLK, which focuses instead on the differences of haplotype frequencies between populations. In contrast to most existing statistics, hapFLK accounts for the hierarchical structure of the sampled populations. Using computer simulations, we show that each of these two features—the use of haplotype information and of the hierarchical structure of populations—significantly improves the detection power of selected loci and that combining them in the hapFLK statistic provides even greater power. We also show that hapFLK is robust with respect to bottlenecks and migration and improves over existing approaches in many situations. Finally, we apply hapFLK to a set of six sheep breeds from Northern Europe and identify seven regions under selection, which include already reported regions but also several new ones. We propose a method to help identifying the population(s) under selection in a detected region, which reveals that in many of these regions selection most likely occurred in more than one population. Furthermore, several of the detected regions correspond to incomplete sweeps, where the favorable haplotype is only at intermediate frequency in the population(s) under selection.     

Screening the pandemic response box identified benzimidazole carbamates,Olorofim and ravuconazole as promising drug candidates for the treatment of eumycetoma

Eumycetoma is a chronic subcutaneous neglected tropical disease that can be caused by more than 40 different fungal causative agents. The most common causative agents produce black grains and belong to the fungal orders Sordariales and Pleosporales. The current antifungal agents used to treat eumycetoma are itraconazole or terbinafine, however, their cure rates are low. To find novel drugs for eumycetoma, we screened 400 diverse drug-like molecules from the Pandemic Response Box against common eumycetoma causative agents as part of the Open Source Mycetoma initiative (MycetOS). 26 compounds were able to inhibit the growth of Madurella mycetomatis, Madurella pseudomycetomatis and Madurella tropicana, 26 compounds inhibited Falciformispora senegalensis and seven inhibited growth of Medicopsis romeroi in vitro. Four compounds were able to inhibit the growth of all five species of fungi tested. They are the benzimidazole carbamates fenbendazole and carbendazim, the 8-aminoquinolone derivative tafenoquine and MMV1578570. Minimal inhibitory concentrations were then determined for the compounds active against M. mycetomatis. Compounds showing potent activity in vitro were further tested in vivo. Fenbendazole, MMV1782387, ravuconazole and olorofim were able to significantly prolong Galleria mellonella larvae survival and are promising candidates to explore in mycetoma treatment and to also serve as scaffolds for medicinal chemistry optimisation in the search for novel antifungals to treat eumycetoma.     

Cobalamin producers and prokaryotic consumers in the Northwest Atlantic

Cobalamin availability can influence primary productivity and ecological interactions in marine microbial communities. The characterization of cobalamin sources and sinks is a first step in investigating cobalamin dynamics and its impact on productivity. Here, we identify potential cobalamin sources and sinks on the Scotian Shelf and Slope in the Northwest Atlantic Ocean. Functional and taxonomic annotation of bulk metagenomic reads, combined with analysis of genome bins, were used to identify potential cobalamin sources and sinks. Cobalamin synthesis potential was mainly attributed to Rhodobacteraceae, Thaumarchaeota, and cyanobacteria (Synechococcus and Prochlorococcus). Cobalamin remodelling potential was mainly attributed to Alteromonadales, Pseudomonadales, Rhizobiales, Oceanospirilalles, Rhodobacteraceae, and Verrucomicrobia, while potential cobalamin consumers include Flavobacteriaceae, Actinobacteria, Porticoccaceae, Methylophiliaceae, and Thermoplasmatota. These complementary approaches identified taxa with the potential to be involved in cobalamin cycling on the Scotian Shelf and revealed genomic information required for further characterization. The Cob operon of Rhodobacterales bacterium HTCC2255, a strain with known importance in cobalamin cycling, was similar to a major cobalamin producer bin, suggesting that a related strain may represent a critical cobalamin source in this region. These results enable future inquiries that will enhance our understanding of how cobalamin shapes microbial interdependencies and productivity in this region.     

Increased level of glycoxidation product N(epsilon)-(carboxymethyl)lysine in rat serum and urine proteins with aging: link with glycoxidative damage accumulation in kidney

Accumulation of carboxymethylated proteins (CML-proteins) is taken as a biomarker of glycoxidative stress which is thought to contribute to the age-related impairment in tissue and cell function. To investigate the occurrence and extent of glycoxidative damage with aging in rat kidney, serum and urine, we have prepared a polyclonal antibody against CML-modified bovine serum albumin. We subsequently used it for immunolocalization and in enzyme-linked immunosorbent assays to evaluate CML-protein content. In the serum, CML-protein level was 1.43+/-0.14 pmol CML/micrograms protein at 3 months and significantly increased by 50% from 10 to 27 months (1.50+/-0.14 pmol CML/micrograms protein vs 2.27+/-0.26 pmol CML/micrograms protein), albumin and transferrin being the main modified proteins. In the urine, CML-protein level was 2.50+/-0.14 pmol CML/micrograms protein at 3 months and markedly increased from 10 months (2.99+/-0.24 pmol CML/micrograms protein) to 27 months (3.76+/-0.25 pmol CML/micrograms protein), with albumin as the main excreted modified protein. Immunolocalization of CML-proteins in kidney provided evidence for an age-dependent increased accumulation in extracellular matrices. Intense staining of the glomerular basement membrane (GBM), Bowman's capsule, and the tubular basement membrane was found. Additionally, the CML content for collagen from GBM was 195.85+/-28.95 pmol CML/microgrms OHPro at 3 months and significantly increased from 10 months (187.61+/-21.99 pmol CML/micrograms OHPro) to 27 months (334.55+/-62.21 pmol CML/micrograms OHPro). These data show that circulating CML-protein level in serum and urine and CML accumulation in nephron extracellular matrices with aging are increasing in parallel. The CML-protein measurement in serum and urine may thus be used as an index for the assessment of age-associated glycoxidative kidney damage.     

"In vivo" intraneuronal trafficking of G protein coupled receptors in the striatum: regulation by dopaminergic and cholinergic environment

We have studied "in vivo" neurochemically identified striatal neurons to analyze the localisation and the trafficking of dopamine and acetylcholine G protein coupled receptors (GPCR) (D1R, D2R, m2R and m4R) under the influence of neurotransmitter environment. We have identified receptors in tissue sections through immunohistochemical detection at the light and electron microscopic level. We have identified receptors in normal animals and after acute and chronic stimulations. We have quantified receptors through image analysis at the electron microscopic level in relation to various subcellular compartments. Our results demonstrate that, in normal conditions, GPCRs are mostly associated with plasma membrane of the striatal neurons, mostly at extra-synaptic sites. In certain instances (m4R; D2R), receptors have prominent localisation inside the rough endoplasmic reticulum. Our results also show that two distinct receptors for a same neurotransmitter may have distinct subcellular localisation in a same neuronal population (m2R versus m4R) and that the same neurotransmitter receptor (m4R) can have distinct localisation in distinct neuronal populations (cytoplasm versus cell surface). After acute stimulation, cell surface receptors undergo dramatic subcellular changes that involve plasma membrane depletion, internalisation in endosomes and in multivesicular bodies. Such changes are reversible after the end of the stimulation and are blocked by antagonist action. Chronic stimulation also provokes changes in subcellular localisation with specific pattern: plasma membrane depletion, and exaggerated storage of receptors in rough endoplasmic reticulum and eventually Golgi complex (D1R; m2R and m4R). Decreasing chronic receptor stimulation reverses such changes. These results demonstrate that, "in vivo", in the striatum, GPCRs undergo complex intraneuronal trafficking under the influence of neurochemical environment in conditions that dramatically modulate the number of cell surface receptors available for interaction with neurotransmitters or drugs. This confirms that "in vivo", the trafficking and the subcellular compartmentalization of GPCRs may contribute to regulate neuronal sensitivity and neuronal interactions in physiological, experimental and pathological conditions, including in therapeutic conditions.     

ProViz: protein interaction visualization and exploration

SUMMARY: ProViz is a tool for the visualization of protein-protein interaction networks, developed by the IntAct European project. It provides facilities for navigating in large graphs and exploring biologically relevant features, and adopts emerging standards such as GO and PSI-MI. AVAILABILITY: ProViz is available under the GPL and may be freely downloaded. Source code and binaries are available at http://cbi.labri.fr/eng/proviz.htm CONTACT: david.sherman@labri.fr