Macromolecular crowding is known to modulate chemical equilibria, reaction rates, and molecular binding events, both in aqueous solutions and at lipid bilayer membranes, natural barriers that enclose the crowded environments of cells and their subcellular compartments. Previous studies on the effects of macromolecular crowding in aqueous compartments on conduction through membranes have focused on single-channel ionic conduction through previously formed pores at thermodynamic equilibrium. Here, the effects of macromolecular crowding on the mechanism of pore formation itself were studied using the droplet interface bilayer (DIB) technique with the voltage-dependent pore-forming peptide alamethicin (alm). Macromolecular crowding was varied using 8 kDa molecular weight polyethylene glycol (PEG8k) or 500 kDa dextran (DEX500k) in two aqueous droplets on both sides of the bilayer membrane. In general, voltage thresholds for pore formation in the presence of crowders in the droplets decreased compared to their values in the absence of crowders, due to excluded volume effects, water binding by PEG, and changes in the ordering of water molecules and hydrogen-bonding interactions involving the polar lipid headgroups. In addition, asymmetric crowder loading (e.g., PEG8k-DEX500k on either side of the membrane) resulted in transmembrane osmotic pressure gradients that either enhanced or degraded the ionic conduction through the pores.
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1021/acs.jpcb.0c01650 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Role Assessment of Water-Soluble Pharmaceutical Form of Phosphatidylcholine on the Catalytic Activity of Cytochrome P450 2C9 and 2D6.
Int J Mol Sci
December 2024
Institute of Biomedical Chemistry, Pogodinskaya Street, 10, Build 8, 119121 Moscow, Russia.
This study aimed to investigate whether the water-soluble pharmaceutical form of phosphatidylcholine nanoparticles (wPC) stimulated the catalytic activity of CYP enzymes 2C9 and 2D6. We have shown that electroenzymatic CYP2C9 catalysis to nonsteroidal anti-inflammatory drug naproxen as a substrate was enhanced from 100% to 155% in the presence of wPC in media. Electroenzymatic CYP2D6 activity in the presence of the adrenoceptor-blocking agent bisoprolol as a substrate was elevated significantly from 100% to 144% when wPC was added to potassium phosphate buffer solution.
View Article and Find Full Text PDFPitfalls of Using ANS Dye Under Molecular Crowding Conditions.
Int J Mol Sci
December 2024
Laboratory of Structural Dynamics, Stability and Folding of Proteins, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Ave., 194064 St. Petersburg, Russia.
The 1-anilino-8-naphthalenesulfonate (ANS) fluorescent dye is widely used in protein folding studies due to the significant increase in its fluorescence quantum yield upon binding to protein hydrophobic regions that become accessible during protein unfolding. However, when modeling cellular macromolecular crowding conditions in protein folding experiments in vitro using crowding agents with guanidine hydrochloride (GdnHCl) as the denaturant, the observed changes in ANS spectral characteristics require careful consideration. This study demonstrates that crowding agents can form clusters that interact differently with ANS.
View Article and Find Full Text PDFDevelopment of Biomimetic Substrates for Limbal Epithelial Stem Cells Using Collagen-Based Films, Hyaluronic Acid, Immortalized Cells, and Macromolecular Crowding.
Life (Basel)
November 2024
Regenerative, Modular & Developmental Engineering Laboratory (REMODEL) and Science Foundation Ireland (SFI) Centre for Research in Medical Devices (CÚRAM), Biomedical Sciences Building, University of Galway, H91 TK33 Galway, Ireland.
Despite the promising potential of cell-based therapies developed using tissue engineering techniques to treat a wide range of diseases, including limbal stem cell deficiency (LSCD), which leads to corneal blindness, their commercialization remains constrained. This is primarily attributable to the limited cell sources, the use of non-standardizable, unscalable, and unsustainable techniques, and the extended manufacturing processes required to produce transplantable tissue-like surrogates. Herein, we present the first demonstration of the potential of a novel approach combining collagen films (CF), hyaluronic acid (HA), human telomerase-immortalized limbal epithelial stem cells (T-LESCs), and macromolecular crowding (MMC) to develop innovative biomimetic substrates for limbal epithelial stem cells (LESCs).
View Article and Find Full Text PDFA cell's global physical state is characterized by its volume and dry mass. The ratio of cell mass to volume is the cell mass density (CMD), which is also a measure of macromolecular crowding and concentrations of all proteins. Using the Fluorescence eXclusion method (FXm) and Quantitative Phase Microscopy (QPM), we investigate CMD dynamics after exposure to sudden media osmolarity change.
View Article and Find Full Text PDFBinding site maturation modulated by molecular density underlies Ndc80 binding to kinetochore receptor CENP-T.
Proc Natl Acad Sci U S A
December 2024
Department of Physiology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104.
Macromolecular assembly depends on tightly regulated pairwise binding interactions that are selectively favored at assembly sites while being disfavored in the soluble phase. This selective control can arise due to molecular density-enhanced binding, as recently found for the kinetochore scaffold protein CENP-T. When clustered, CENP-T recruits markedly more Ndc80 complexes than its monomeric counterpart, but the underlying molecular basis remains elusive.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!