Journal of Membrane Biology最新文献

Fluoxetine (FLX), used in the clinic to treat depression, is a well-known cationic amphiphilic antidepressant. However, there is a lack of research on the effect of FLX on the surface behavior of lipid monolayers under different surface pressures. In this study, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/CHOL (DPPC/POPC/CHOL) monolayers were prepared via the Langmuir method, and FLX was added to these monolayers under various surface pressures. The effect of FLX on the surface behavior of DPPC/POPC/CHOL monolayers under various surface pressures was studied using a combination of surface pressure-area isotherms, compressibility modulus-surface pressure curves, and atomic force microscope (AFM). The results showed that the effect of FLX on the lipid monolayers was different under different surface pressures. The interaction between FLX and lipid molecules was weak under low surface pressures, and FLX could easily intercalate between the lipid molecules to inhibit monolayer phase transition. The interaction between FLX and lipid molecules was enhanced and FLX tended to self-aggregate to reduce the monolayer stability when the surface pressure was high. This study lays the foundation for further studies on the interaction between FLX and lipid monolayers.

Epithelial barriers constitute a fundamental requirement in every organism, as they allow the separation of different environments and set boundaries against noxious and other adverse effectors. In many inflammatory and degenerative diseases, epithelial barrier function is impaired because of a disturbance of the paracellular seal. Recently, the Xenopus laevis oocyte has been established as a heterologous expression model for the analysis of transmembrane tight junction protein interactions and is currently considered to be a suitable screening model for barrier effectors. A prerequisite for this application is a physiological anchoring of claudins to the cytoskeleton via the major scaffolding protein tjp1 (tight junction protein 1, ZO-1). We have analyzed the oocyte model with regard to the interaction of heterologously expressed claudins and tjp1. Our experiments have revealed endogenous tjp1 expression in protein and mRNA analyses of unfertilized Xenopus laevis oocytes expressing human claudin 1 (CLDN1) to claudin 5 (CLDN5). The amphibian cell model can therefore be used for the analysis of claudin interactions.

Shielding of the specific body organ using the biocompatible material helps preventing direct exposure of that part to the foreign entities responsible for infections. Here we show the potential of the A2 milk protein recovered from the milk of cow from Indian origin for possible prevention of the direct exposure to other foreign molecules. We measured the surface pressure of the monolayers of different types of protein samples using Langmuir isotherm experiments. The surface pressure measurements for the monolayer of four types of protein macromolecules have been carried out using the Wilhelmy plate micro pressure sensor. We studied the self-organization of different protein macromolecules and their monolayer compression characteristics. The electrochemical behaviour is studied using electrochemical impedance spectroscopy. We found the highest surface pressure for the monolayer of A2 protein. Further, it is also found that A2 protein exhibited the highest surface activity amongst the other proteins. This property can be effectively used for making the envelope of the A2 protein surrounding the targeted entity.

Aquaporins constitute a family of transmembrane proteins that function to transport water and other small solutes across the cell membrane. Aquaporins family members are found in diverse life forms. Aquaporins share the common structural fold consisting of six transmembrane alpha helices with a central water-transporting channel. Four such monomers assemble together to form tetramers as their biological unit. Initially, aquaporins were discovered as water-transporting channels, but several studies supported their involvement in mediating the facilitated diffusion of different solutes. The so-called water channel is able to transport a variety of substrates ranging from a neutral molecule to a charged molecule or a small molecule to a bulky molecule or even a gas molecule. This article gives an overview of a diverse range of substrates conducted by aquaporin family members. Prime focus is on human aquaporins where aquaporins show a wide tissue distribution and substrate specificity leading to various physiological functions. This review also highlights the structural mechanisms leading to the transport of water and glycerol. More research is needed to understand how one common fold enables the aquaporins to transport an array of solutes.

Exosomes are special extracellular vesicles secreted by cells, which are of great significance in the basic research of life science and clinical application and has become a hot research field with rapid development in recent 10 years. Therefore, the isolation and separation of exosomes is particularly important for the research and application of exosomes. This paper aims to review the research progress of exosome isolation and separation methods in recent years, including ultracentrifugation, ultrafiltration, size‑exclusion chromatography, precipitation, immunomagnetic bead capture method, aptamer-based isolation, and isolation methods based on microfluidic technology. It is generally accepted that most of the existing methods have limitations, for example, ultracentrifugation is time-consuming and laborious, and immunomagnetic bead capture method and aptamer-based separation method have small sample processing capacity and high cost. As a result, we also introduce some common situations in which two or more methods are combined for use. Finally, the separation and isolation methods including all those presented in this review were compared and summarized.

Most blockers of both hERG (human ether-à-go-go-related gene) channels and pancreatic ß-cell ATP-sensitive K⁺ (K_ATP) channels access their binding sites from the cytoplasmic side of the plasma membrane. It is unknown whether binding to intracellular components competes with binding of these substances to K⁺ channels. The whole-cell configuration of the patch-clamp technique, a laser-scanning confocal microscope, and fluorescence correlation spectroscopy (FCS) were used to study hERG channels expressed in HEK (human embryonic kidney) 293 cells and K_ATP channels from the clonal insulinoma cell line RINm5F. When applied via the pipette solution in the whole-cell configuration, terfenadine blocked both hERG and K_ATP currents with much lower potency than after application via the bath solution, which was not due to P-glycoprotein-mediated efflux of terfenadine. Such a difference was not observed with dofetilide and tolbutamide. 37-68% of hERG/EGFP (enhanced green-fluorescent protein) fusion proteins expressed in HEK 293 cells were slowly diffusible as determined by laser-scanning microscopy in the whole-cell configuration and by FCS in intact cells. Bath application of a green-fluorescent sulphonylurea derivative (Bodipy-glibenclamide) induced a diffuse fluorescence in the cytosol of RINm5F cells under whole-cell patch-clamp conditions. These observations demonstrate the presence of intracellular binding sites for hERG and K_ATP channel blockers not dialyzable by the patch-pipette solution. Intracellular binding of terfenadine was not influenced by a mutated hERG (Y652A) channel. In conclusion, substances with high lipophilicity are not freely diffusible inside the cell but steep concentration gradients might exist within the cell and in the sub-membrane space.

Pore-forming proteins (PFPs) are produced by various organisms, including pathogenic bacteria, and form pores within the target cell membrane. Streptolysin O (SLO) is a PFP produced by Streptococcus pyogenes and forms high-order oligomers on the membrane surface. In this prepore state, multiple α-helices in domain 3 of each subunit exist as unfolded structures and transiently interact with each other. They subsequently transition into transmembrane β-hairpins (TMHs) and form pores with diameters of 20-30 nm. However, in this pore formation process, the trigger of the transition in a subunit and collaboration between subunits remains elusive. Here, I observed the dynamic pore formation process using high-speed atomic force microscopy. During the oligomer transition process, each subunit was sequentially inserted into the membrane, propagating along the oligomer in a domino-like fashion (chain reaction). This process also occurred on hybrid oligomers containing wildtype and mutant subunits, which cannot insert into the membrane because of an introduced disulfide bond. Furthermore, propagation still occurred when an excessive force was added to hybrid oligomers in the prepore state. Based on the observed chain reactions, I estimate the free energies and forces that trigger the transition in a subunit. Furthermore, I hypothesize that the collaboration between subunits is related to the structure of their TMH regions and interactions between TMH-TMH and TMH-lipid molecules.