Biology subjects

Vazdar, M.

Publications and source records attributed to Vazdar, M..

3 recordsLinked to original sources

bioRxiv2023

A Stealthy Player in Lipid Experiments? EDTA Binding to Phosphatidylcholine Membranes Probed by Simulations and Monolayer Experiments

Ethylenediaminetetraacetic acid (EDTA) is frequently used in lipid experiments to remove redundant ions, such as Ca2+, from the sample solution. In this work, combining molecular dynamics (MD) simulations and Langmuir monolayer experiments, we show that on top of the expected Ca2+ depletion, EDTA anions themselves bind to phosphatidylcholine (PC) monolayers. This binding, originating from EDTA interaction with choline groups of PC lipids, leads to the adsorption of EDTA anions at the monolayer surface and concentrationdependent changes in surface pressure as measured by monolayer experiments and explained by MD simulations. This surprising observation emphasizes that lipid experiments carried out using EDTA-containing solutions, especially of high concentrations, must be interpreted very carefully due to potential interfering interactions of EDTA with lipids and other biomolecules involved in the experiment, e.g., cationic peptides, that may alter membranebinding affinities of studied compounds. TOC Figure O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=135 SRC="FIGDIR/small/532294v1_ufig1.gif" ALT="Figure 1"> View larger version (80K): org.highwire.dtl.DTLVardef@128c201org.highwire.dtl.DTLVardef@d8eaeborg.highwire.dtl.DTLVardef@126edd0org.highwire.dtl.DTLVardef@c6f868_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2022

Membrane lipid reshaping underlies oxidative stress sensing by the mitochondrial proteins UCP1 and ANT1

Oxidative stress and ROS are important players in the pathogenesis of several diseases. Besides the direct modification of proteins, ROS modify lipids with negative spontaneous curvature, such as phosphatidylethanolamine (PE), producing PE adducts and lysolipids. The formation of PE-adducts potentiates the protonophoric activity of the uncoupling protein 1 (UCP1), but the molecular mechanism remains obscure. Here, we connected the ROS-mediated lipid shape alteration with the membrane mechanical properties and function of UCP1 and adenine nucleotide translocase 1 (ANT1). We found that lysophosphatidylcholines (OPC and MPC) and PE adducts decrease a bending modulus in lipid bilayers and increase the protonophoric activity of both proteins. Furthermore, MD simulations revealed that modified PEs and lysolipids alter the membrane lateral pressure profile in the same direction and range, indicating that modified PEs act as lipids with positive spontaneous curvature. Both results indicate that oxidative stress decreases stored curvature elastic stress (SCES) in the lipid bilayer membrane. In conclusion, we demonstrate that UCP1 and ANT1 sense SCES and propose a new regulatory mechanism for the function of these proteins.

biophysics↗

bioRxiv2022

Mechanism of the ANT-mediated transport of fatty acid anions across the inner mitochondrial membrane

The additional protonophoric function of the mitochondrial adenine nucleotide translocase (ANT1) is now recognized. However, the molecular mechanism remains controversial. Fatty acid (FA) cycling hypothesis postulates that FAs transport protons across the inner mitochondrial membrane to the matrix by a flip-flop, whereas ANT1 facilitates the translocation of FA anions (FA-) back to the intermembrane space. By a combined approach involving measurements of current through the planar lipid bilayers reconstituted with recombinant ANT1, site-directed mutagenesis and molecular dynamics simulations, we show that FA- is initially caught by R59 on the matrix side of ANT1, then moves along the positively charged protein-lipid interface, and binds to R79, where it is protonated in the hydrated cavity in the presence of D134. R79 is crucial for the competitive binding of ANT1 substrates (ATP and ADP) and inhibitors (carboxyatractyloside, bongkrekic acid). The binding sites are well-conserved in mitochondrial SLC25 members, implying a general transporting mechanism for FA anions.

biophysics↗