Changes in single K+ channel behavior through the lipid phase transition

We show that the activity of an ion channel is strictly related to the phase state of the lipid bilayer hosting the channel. By measuring unitary conductance, dwell times, and open probability of the K+ channel KcsA as a function of temperature in lipid bilayers composed of POPE and POPG in different relative proportions, we obtain that all those properties show a trend inversion when the bilayer is in the transition region between the liquid disordered and the solid ordered phase. These data suggest that the physical properties of the lipid bilayer influence ion channel activity likely via a fine tuning of its conformations. In a more general interpretative framework, we suggest that other parameters such as pH, ionic strength, and the action of amphiphilic drugs can affect the physical behavior of the lipid bilayer in a fashion similar to temperature changes resulting in functional changes of transmembrane proteins

Dynamic Force Spectroscopy on Supported Lipid Bilayers: Effect of Temperature and Sample Preparation

AbstractBiological membranes are constantly exposed to forces. The stress-strain relation in membranes determines the behavior of many integral membrane proteins or other membrane related-proteins that show a mechanosensitive behavior. Here, we studied by force spectroscopy the behavior of supported lipid bilayers (SLBs) subjected to forces perpendicular to their plane. We measured the lipid bilayer mechanical properties and the force required for the punch-through event characteristic of atomic force spectroscopy on SLBs as a function of the interleaflet coupling. We found that for an uncoupled bilayer, the overall tip penetration occurs sequentially through the two leaflets, giving rise to two penetration events. In the case of a bilayer with coupled leaflets, penetration of the atomic force microscope tip always occurred in a single step. Considering the dependence of the jump-through force value on the tip speed, we also studied the process in the context of dynamic force spectroscopy (DFS). We performed DFS experiments by changing the temperature and cantilever spring constant, and analyzed the results in the context of the developed theories for DFS. We found that experiments performed at different temperatures and with different cantilever spring constants enabled a more effective comparison of experimental data with theory in comparison with previously published data

Supported lipid bilayers on mica and silicon oxide: comparison of the main phase transition behavior

The usual biophysical approach to the study of biological membranes is that of turning to model systems. From these models, general physical principles ruling the lateral membrane structure can be obtained. A promising model system is the supported lipid bilayer (SLB) which could foresee the simultaneous investigation of the structure and physical properties of lipid bilayers reconstituted with membrane proteins. A complete exploitation of the model system to retrieve biologically relevant information requires an in-depth knowledge of the possible effect that experimental parameters could have on the behavior of the SLB. Here we used atomic force microscopy (AFM) to study the effect of different types of substrates on the behavior of SLBs as far as their main phase transition is concerned. We found that different substrates (mica and silicon oxide) can affect in dissimilar ways the interleaflet coupling of the bilayer, which might represent a sort of lipid signaling allowing communication between receptors on the extracellular leaflet and cytoplasmic components. By decreasing the interaction between the SLB and the substrate the interleaflet coupling is preserved independently of the bilayer preparation strategy. Moreover, we investigated by time-lapse AFM an isothermal phase transition induced by a pH change on a SLB. We established that the presence of a pH gradient across the bilayer can weaken the strength of the interleaflet coupling which is present in symmetrical pH conditions

Supported lipid bilayers on mica and silicon oxide: comparison of the main phase transition behavior

The usual biophysical approach to the study of biological membranes is that of turning to model systems. From these models, general physical principles ruling the lateral membrane structure can be obtained. A promising model system is the supported lipid bilayer (SLB) which could foresee the simultaneous investigation of the structure and physical properties of lipid bilayers reconstituted with membrane proteins. A complete exploitation of the model system to retrieve biologically relevant information requires an in-depth knowledge of the possible effect that experimental parameters could have on the behavior of the SLB. Here we used atomic force microscopy (AFM) to study the effect of different types of substrates on the behavior of SLBs as far as their main phase transition is concerned. We found that different substrates (mica and silicon oxide) can affect in dissimilar ways the interleaflet coupling of the bilayer, which might represent a sort of lipid signaling allowing communication between receptors on the extracellular leaflet and cytoplasmic components. By decreasing the interaction between the SLB and the substrate the interleaflet coupling is preserved independently of the bilayer preparation strategy. Moreover, we investigated by time-lapse AFM an isothermal phase transition induced by a pH change on a SLB. We established that the presence of a pH gradient across the bilayer can weaken the strength of the interleaflet coupling which is present in symmetrical pH conditions

What do we really measure in AFM punch-through experiments on supported lipid bilayers?

Nowadays, there is much experimental evidence that the mechanical properties of biological membranes affect membrane protein functions. A very convenient technique to study these properties on a spatial scale relevant to that of single proteins is represented by Atomic Force Spectroscopy (AFS). In this study we measured the force the AFM tip has to apply on a supported lipid bilayer to punch-through it as a function of different environmental parameters. We observed that this force is reduced when the lipid bilayer is in its phase transition region. We interpreted our results on the basis of thermodynamical considerations and we stressed their biological relevance. In particular, the reduced punch-through force in the phase transition region could be relevant for the function of membrane proteins which operates by conformational changes at the protein/lipid interface. We also suggest that the presence of a transmembrane voltage drop can affect the measured punch-through force

What do we really measure in AFM punch-through experiments on supported lipid bilayers?

Nowadays, there is much experimental evidence that the mechanical properties of biological membranes affect membrane protein functions. A very convenient technique to study these properties on a spatial scale relevant to that of single proteins is represented by Atomic Force Spectroscopy (AFS). In this study we measured the force the AFM tip has to apply on a supported lipid bilayer to punch-through it as a function of different environmental parameters. We observed that this force is reduced when the lipid bilayer is in its phase transition region. We interpreted our results on the basis of thermodynamical considerations and we stressed their biological relevance. In particular, the reduced punch-through force in the phase transition region could be relevant for the function of membrane proteins which operates by conformational changes at the protein/lipid interface. We also suggest that the presence of a transmembrane voltage drop can affect the measured punch-through force

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