Impact of Physico-Chemical Properties of Cellulose Nanocrystal/Silver Nanoparticle Hybrid Suspensions on Their Biocidal and Toxicological Effects.

There is a demand for nanoparticles that are environmentally acceptable, but simultaneously efficient and low cost. We prepared silver nanoparticles (AgNPs) grafted on a native bio-based substrate (cellulose nanocrystals, CNCs) with high biocidal activity and no toxicological impact. AgNPs of 10 nm are nucleated on CNCs in aqueous suspension with content from 0.

ZnO and TiO2 nanoparticles alter the ability of Bacillus subtilis to fight against a stress.

Due to the physicochemical properties of nanoparticles, the use of nanomaterials increases over time in industrial and medical processes. We herein report the negative impact of nanoparticles, using solid growth conditions mimicking a biofilm, on the ability of Bacillus subtilis to fight against a stress. Bacteria have been exposed to sublethal doses of nanoparticles corresponding to conditions that bacteria may meet in their natural biotopes, the upper layer of soil or the gut microbiome.

Author Correction: Impact of nanoparticles on the Bacillus subtilis (3610) competence.

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Impact of nanoparticles on the Bacillus subtilis (3610) competence.

Due to the physicochemical properties of nanoparticles, the use of nanomaterials increases every year in industrial and medical processes. At the same time, the increasing number of bacteria becoming resistant to many antibiotics, mostly by a horizontal gene transfer process, is a major public health concern. We herein report, for the first time, the role of nanoparticles in the physiological induction of horizontal gene transfer in bacteria.

Zinc oxide induces the stringent response and major reorientations in the central metabolism of Bacillus subtilis.

Microorganisms, such as bacteria, are one of the first targets of nanoparticles in the environment. In this study, we tested the effect of two nanoparticles, ZnO and TiO2, with the salt ZnSO4 as the control, on the Gram-positive bacterium Bacillus subtilis by 2D gel electrophoresis-based proteomics. Despite a significant effect on viability (LD50), TiO2 NPs had no detectable effect on the proteomic pattern, while ZnO NPs and ZnSO4 significantly modified B.

Functional consequences of the over-expression of TRPC6 channels in HEK cells: impact on the homeostasis of zinc.

The canonical transient receptor potential 6 (TRPC6) protein is a non-selective cation channel able to transport essential trace elements like iron (Fe) and zinc (Zn) through the plasma membrane. Its over-expression in HEK-293 cells causes an intracellular accumulation of Zn, indicating that it could be involved in Zn transport. This finding prompted us to better understand the role played by TRPC6 in Zn homeostasis.

Improved proteomic analysis of nuclear proteins, as exemplified by the comparison of two myeloid cell lines nuclear proteomes.

One of the challenges of the proteomic analysis by 2D-gel is to visualize the low abundance proteins, particularly those localized in the organelles. An additional problem with nuclear proteins lies in their strong interaction with nuclear acids. Several experimental procedures have been tested to increase, in the nuclear extract, the ratio of nuclear proteins compared to contaminant proteins, and also to obtain reproducible conditions compatible with 2D-gel electrophoresis.

Improvements and simplifications in in-gel fluorescent detection of proteins using ruthenium II tris-(bathophenanthroline disulfonate): the poor man's fluorescent detection method.

Fluorescent detection of proteins is a popular method of detection allying sensitivity, linearity and compatibility with mass spectrometry. Among the numerous methods described in the literature, staining with ruthenium II tris(bathophenanthroline disulfonate) is particularly cost-effective, but slightly cumbersome owing to difficulties in the preparation of the complex and complexity of staining protocols. We describe here the modifications on both aspects that allow to perform a higher contrast staining and offer a more robust method of complex preparation, thereby maximizing the advantages of the method.

Two-dimensional gel electrophoresis in proteomics: Past, present and future.

Two-dimensional gel electrophoresis has been instrumental in the birth and developments of proteomics, although it is no longer the exclusive separation tool used in the field of proteomics. In this review, a historical perspective is made, starting from the days where two-dimensional gels were used and the word proteomics did not even exist. The events that have led to the birth of proteomics are also recalled, ending with a description of the now well-known limitations of two-dimensional gels in proteomics.

Silver staining of proteins in 2DE gels.

Silver staining detects proteins after electrophoretic separation on polyacrylamide gels. Its main positive features are its excellent sensitivity (in the low nanogram range) and the use of very simple and cheap equipment and chemicals. The sequential phases of silver staining are protein fixation, then sensitization, then silver impregnation, and finally image development.

Sweet silver: a formaldehyde-free silver staining using aldoses as developing agents, with enhanced compatibility with mass spectrometry.

Protein detection methods after electrophoresis have to be sensitive, homogeneous, and not to impair downstream analysis of proteins by MS. Speed, low cost, and user friendliness are also favored features. Silver staining combines many of these features, but its compatibility with MS is limited.

Fully denaturing two-dimensional electrophoresis of membrane proteins: a critical update.

The quality and ease of proteomics analysis depends on the performance of the analytical tools used, and thus of the performances of the protein separation tools used to deconvolute complex protein samples. Among protein samples, membrane proteins are one of the most difficult sample classes, because of their hydrophobicity and embedment in the lipid bilayers. This review deals with the recent progresses and advances made in the separation of membrane proteins by 2-DE separating only denatured proteins.

Organelle proteomics.

This unit describes strategies for studying the proteomes of organelles, which is one example of targeted proteomics. It relies heavily on previously published units dealing with organelle preparation, protein solubilization, and proteomics techniques. A specific commentary for organelle proteomics is provided.

Preparing protein extracts for quantitative two-dimensional gel comparison.

This unit describes basic protocols for efficient and reproducible protein solubilization from a variety of biological samples, including cultured animal cells and tissues, plant cells and tissues, bacteria, nuclei, other subcellular organelles, plasma, serum, and other biological fluids. The optimized extraction process is strongly sample-dependent and cannot be described for every type of sample. Instead, typical protocols are provided as general guidelines and illustrate good starting points for sample-preparation optimization.

Purification and mass spectrometry identification of microtubule-binding proteins from Xenopus egg extracts.

Microtubule-binding proteins are conveniently divided into two large groups: MAPs (microtubule-associated proteins), which can stabilize, anchor, and/or nucleate microtubules, and motors, which use the energy of ATP hydrolysis for a variety of functions, including microtubule network organization and cargo transportation along microtubules. Here, we describe the use of Taxol-stabilized microtubules for purification of MAPs, motors, and their complexes from Xenopus egg extracts. Isolated proteins are analysed using sodium dodecyl sulfate gel electrophoresis and identified by various mass spectrometry and database mining technologies.

Ultrafast coelectrophoretic fluorescent staining of proteins with carbocyanines.

Protein detection on SDS gels or on 2-D gels must combine several features, such as sensitivity, homogeneity from one protein to another, speed, low cost, and user-friendliness. For some applications, it is also interesting to have a nonfixing stain, so that proteins can be mobilized from the gel for further use (electroelution, blotting). We show here that coelectrophoretic staining by fluorophores of the oxacarbocyanine family, and especially diheptyloxacarbocyanine, offers several positive features.

Silver staining of proteins in polyacrylamide gels.

Silver staining is used to detect proteins after electrophoretic separation on polyacrylamide gels. It combines excellent sensitivity (in the low nanogram range) with the use of very simple and cheap equipment and chemicals. It is compatible with downstream processing, such as mass spectrometry analysis after protein digestion.

The Crl-RpoS regulon of Escherichia coli.

The RpoS subunit of RNA polymerase controls the expression of numerous genes involved in stationary phase and in response to different stress conditions. The regulatory protein Crl increases the activity of RpoS by direct interaction with the RpoS holoenzyme. To define the extent of the Crl regulon, we used two-dimensional SDS-PAGE to measure the role of Crl in regulating the expression of the Escherichia coliproteome in stationary phase at 30 degrees C.

Detergents and chaotropes for protein solubilization before two-dimensional electrophoresis.

Because of the outstanding separating capabilities of two-dimensional electrophoresis for complete proteins, it would be advantageous to be able to apply it to all types of proteins. Unfortunately, severe solubility problems hamper the analysis of many classes of proteins, but especially membrane proteins. These problems arise mainly in the extraction and isoelectric focusing steps, and solutions are sought to improve protein solubility under the conditions prevailing during isoelectric focusing.

Protein arginylation in rat brain cytosol: a proteomic analysis.

Arginine can be post-translationally incorporated from arginyl-tRNA into the N-terminus of soluble acceptor proteins in a reaction catalyzed by arginyl-tRNA protein transferase. In the present study, several soluble rat brain proteins that accepted arginine were identified after arginine incorporation by two dimensional electrophoresis and mass spectrometry. They were identified as: contrapsin-like protease inhibitor-3, alpha-1-antitrypsin, apolipoprotein E, hemopexin, calreticulin and apolipoprotein A-I.

Improved mass spectrometry compatibility is afforded by ammoniacal silver staining.

Sequence coverage in MS analysis of protein digestion-derived peptides is a key issue for detailed characterization of proteins or identification at low quantities. In gel-based proteomics studies, the sequence coverage greatly depends on the protein detection method. It is shown here that ammoniacal silver detection methods offer improved sequence coverage over standard silver nitrate methods, while keeping the high sensitivity of silver staining.

High expression of antioxidant proteins in dendritic cells: possible implications in atherosclerosis.

Dendritic cells (DCs) display the unique ability to activate naive T cells and to initiate primary T cell responses revealed in DC-T cell alloreactions. DCs frequently operate under stress conditions. Oxidative stress enhances the production of inflammatory cytokines by DCs.

Oxidative stress response: a proteomic view.

The oxidative stress response is characterized by various effects on a range of biologic molecules. When examined at the protein level, both expression levels and protein modifications are altered by oxidative stress. While these effects have been studied in the past by classic biochemical methods, the recent onset of proteomics methods has allowed the oxidative stress response to be studied on a much wider scale.

FMRP interferes with the Rac1 pathway and controls actin cytoskeleton dynamics in murine fibroblasts.

Fragile X syndrome, the most common form of inherited mental retardation, is caused by absence of FMRP, an RNA-binding protein implicated in regulation of mRNA translation and/or transport. We have previously shown that dFMR1, the Drosophila ortholog of FMRP, is genetically linked to the dRac1 GTPase, a key player in actin cytoskeleton remodeling. Here, we demonstrate that FMRP and the Rac1 pathway are connected in a model of murine fibroblasts.

About the mechanism of interference of silver staining with peptide mass spectrometry.

The mechanism by which silver staining of proteins in polyacrylamide gels interferes with mass spectrometry of peptides produced by proteolysis has been investigated. It was demonstrated that this interference increases with time between silver staining and gel processing, although the silver image is constant. This suggested an important role of the formaldehyde used in silver staining development in this interference process.