Small-Angle Neutron Scattering Reveals the Nanostructure of Liposomes with Embedded OprF Porins of .

The use of liposomes as drug delivery systems emerged in the last decades in view of their capacity and versatility to deliver a variety of therapeutic agents. By means of small-angle neutron scattering (SANS), we performed a detailed characterization of liposomes containing outer membrane protein F (OprF), the main porin of the bacterium outer membrane. These OprF-liposomes are the basis of a novel vaccine against this antibiotic-resistant bacterium, which is one of the main hospital-acquired pathogens and causes each year a significant number of deaths.

Cell-free expression of the outer membrane protein OprF of for vaccine purposes.

is the second-leading cause of nosocomial infections and pneumonia in hospitals. Because of its extraordinary capacity for developing resistance to antibiotics, treating infections by is becoming a challenge, lengthening hospital stays, and increasing medical costs and mortality. The outer membrane protein OprF is a well-conserved and immunogenic porin playing an important role in quorum sensing and in biofilm formation.

A PANI supported lipid bilayer that contains NhaA transporter proteins provides a basis for a biomimetic biocapacitor.

We designed a supported lipid bilayer (SLB) biomimetic membrane system that comprised polyaniline (PANI) to support a lipid bilayer membrane that incorporated Na+/H+ transporter proteins (NhaA) to give the system the capability of controllable electrogenic ion transport. The high turnover rate of NhaA (∼105 per min) provides the basis for this PANI-SLB-NhaA system to be a high-speed rechargeable biocapacitor that functions as a low-energy-consuming fast switch for biological engineering applications.

Functional Characterization of Cell-Free Expressed OprF Porin from Pseudomonas aeruginosa Stably Incorporated in Tethered Lipid Bilayers.

OprF has a central role in Pseudomonas aeruginosa virulence and thus provides a putative target for either vaccines or antibiotic cofactors that could overcome the bacterium's natural resistance to antibiotics. Here we describe a procedure to optimize the production of highly pure and functional OprF porins that are then incorporated into a tethered lipid bilayer. This is a stable biomimetic system that provides the capability to investigate structural aspects and function of OprF using and neutron reflectometry and electrical impedance spectroscopy.

[Biomimetic sensors in biomedical research].

The recent research on both the synthesis of membrane proteins by cell-free systems and the reconstruction of planar lipid membranes, has led to the development of a cross-technology to produce biosensors or filters. Numerous biomimetic membranes are currently being standardized and used by the industry, such as filters containing aquaporin for water desalination, or used in routine at the laboratory scale, for example the bacteriorhodopsin as a light sensor. In the medical area, several fields of application of these biomimetic membranes are under consideration today, particularly for the screening of therapeutic molecules and for the developing of new tools in diagnosis, patient monitoring and personalized medicine.

A common highly conserved cadmium detoxification mechanism from bacteria to humans: heavy metal tolerance conferred by the ATP-binding cassette (ABC) transporter SpHMT1 requires glutathione but not metal-chelating phytochelatin peptides.

Cadmium poses a significant threat to human health due to its toxicity. In mammals and in bakers' yeast, cadmium is detoxified by ATP-binding cassette transporters after conjugation to glutathione. In fission yeast, phytochelatins constitute the co-substrate with cadmium for the transporter SpHMT1.

Transport of antimony salts by Arabidopsis thaliana protoplasts over-expressing the human multidrug resistance-associated protein 1 (MRP1/ABCC1).

ABC transporters from the multidrug resistance-associated protein (MRP) subfamily are glutathione S-conjugate pumps exhibiting a broad substrate specificity illustrated by numerous xenobiotics, such as anticancer drugs, herbicides, pesticides and heavy metals. The engineering of MRP transporters into plants might be interesting either to reduce the quantity of xenobiotics taken up by the plant in the context of "safe-food" strategies or, conversely, in the development of phytoremediation strategies in which xenobiotics are sequestered in the vacuolar compartment. In this report, we obtained Arabidopsis transgenic plants overexpressing human MRP1.

Characterization of the typical multidrug resistance profile in human uveal melanoma cell lines and in mouse liver metastasis derivatives.

Uveal melanoma is the most common intraocular malignancy. To study its biology, stable cell lines provide a useful tool, but these are very difficult to obtain. A stable and rapidly growing human choroidal melanoma cell line composed of pure epithelioid cells was established and maintained for at least 4 years.

Major cytogenetic aberrations and typical multidrug resistance phenotype of uveal melanoma: current views and new therapeutic prospects.

Uveal melanoma is the most frequent intra-ocular cancer. The recent development of new chromosome-related technologies have permitted the elucidation of both the cytogenetics and the natural history of this disease. Fifty to 60% of uveal melanomas are linked to a monosomy 3, which appears as an early and determinant event in tumor progression.

Control of P-glycoprotein activity by membrane cholesterol amounts and their relation to multidrug resistance in human CEM leukemia cells.

P-glycoprotein (P-gp) is the most well-known ATP-binding cassette (ABC) transporter involved in unidirectional substrate translocation across the membrane lipid bilayer, thereby causing the typical multidrug resistance (MDR) phenotype expressed in many cancers. We observed that in human CEM acute lymphoblastic leukemia cells expressing various degrees of chemoresistance and where P-gp was the sole MDR-related ABC transporter detected, the amount of esterified cholesterol increased linearly with the level of resistance to vinblastine while the amounts of total and free cholesterol increased in a nonlinear way. Membrane cholesterol controlled the ATPase activity of P-gp in a linear manner, whereas the P-gp-induced daunomycin efflux decreased nonlinearly with the depletion of membrane cholesterol.

Multidrug-resistant cancer cells contain two populations of P-glycoprotein with differently stimulated P-gp ATPase activities: evidence from atomic force microscopy and biochemical analysis.

Considerable interest exists about the localization of P-gp (P-glycoprotein) in DRMs (detergent-resistant membranes) of multidrug resistant cancer cells, in particular concerning the potential modulating role of the closely related lipids and proteins on P-gp activity. Our observation of the opposite effect of verapamil on P-gp ATPase activity from DRM and solubilized-membrane fractions of CEM-resistant leukaemia cells, and results from Langmuir experiments on membrane monolayers from resistant CEM cells, strongly suggest that two functional populations of P-gp exist. The first is located in DRM regions: it displays its optimal P-gp ATPase activity, which is almost completely inhibited by orthovanadate and activated by verapamil.

New invMED1 element cis-activates human multidrug-related MDR1 and MVP genes, involving the LRP130 protein.

The MDR1 gene is a key component of the cytotoxic defense network and its overexpression results in the multidrug resistance (MDR) phenotype. However, the molecular mechanisms that regulate the MDR1 gene and coordinate multiple MDR-related genes expression are poorly understood. In a previous study, we identified a new 12 bp cis-activating region in the 5'-flanking region of the human MDR1 gene, which we called inverted MED1.

Transcriptional regulation of the human MDR1 gene at the level of the inverted MED-1 promoter region.

The typical multidrug resistance phenotype (MDR), the major cause of failure of cancer chemotherapy, is the result of the overexpression of the human MDR1 gene, the regulation of which is still incompletely understood. Using several EMSA experiments, we have identified a new regulatory sequence located from -103 to -98 bp relative to the +1 start site in the MDR1 promoter region. This sequence, which we called inverted MED-1, acts as a cis-activator for this gene.

Transcriptional regulators of the human multidrug resistance 1 gene: recent views.

The multidrug resistance (MDR) phenotype is the major cause of failure of cancer chemotherapy. This phenotype is mainly due to the overexpression of the human MDR1 (hMDR1) gene. Several studies have shown that transcriptional regulation of this gene is unexpectedly complex and is far from being completely understood.