Final Report Summary - NANOORDER (Nanoscale Organisation and Dynamics of Ionic Networks at Biological Interfaces)
In solution, ions tend to accumulate near charged surfaces and form an electrostatic double layer. The ionic density of the double layer is generally well understood, but little is known about the lateral organization of the ions at the surface. The existence of ordered ionic networks could have dramatic consequences for interfacial processes, for example affecting the mechanical properties of biomembranes and help re-shape the cell. Recent developments in the field of atomic force microscopy (AFM) have made it possible to image single ions on surfaces and study the formation of organized networks over tens of nanometres with molecular precision. This project investigates the formation of such structures at model bio-interfaces, in solution. The aim is to unravel the mechanisms that allow soft bio-membranes to control interfacial ionic organisation through their hydration and physical properties.
The project is organised along 4 main research lines:
1. Formation of ionic networks at the surface of gel-phase lipid bilayers.
The existence of nanoscale ionic networks has never been observed at the surface of lipid membranes. Using ‘static’ gel-phase membrane offer an ideal starting point to showing the existence of possible networks and investigate their properties.
Several important results have stemmed from the project. Arguably the main result is the confirmation of postulated long-lived ionic networks at the surface of charged membranes. Atomic-level images in solution show that the networks strongly depend on the types of ions involved both in size and stability. These ionic networks evolve remarkably slowly, typically over seconds to minutes. Computer simulations showed that these mesoscale clusters are ion-specific and their cohesion originates from water-mediated interfaces involving the adsorbed ions in different hydration states and the lipid headgroups. We demonstrate the networks to locally change the stiffness of the membrane by more than 10%, effectively providing a mechanism for membranes to passively modulate their mechanical properties at the nanoscale in a spontaneous manner (without added external energy).
These results are unprecedented and suggest that our current understanding of biological membranes misses an important regulation mechanism. Our current understanding of the regulation of membrane shape, mechanical and viscous properties involve specialized proteins and molecules. This project demonstrates that interfacial effects can be sufficient to achieve similar results, and therefore need to be considered in any comprehensive model of biomembranes. We foresee the results to have a direct impact on both fundamental science and socio-economical applications. First the novelty and groundbreaking nature of our results is expected to lead to several significant scientific publications and presentations in conferences and meeting. The results are also expected to change our current understanding of biological membranes by including interfacial effects as an integral part of the membrane. We expect this view to eventually find its way into teaching manuals and become part of the standard model accepted by students and specialists alike. Second, the results will have implications for the pharmaceutical industry, in particular for the development and testing of new drugs that can exploit the reported ionic networks.
2. Impact of ions on the behaviour of fluid lipid membranes.
In most natural systems, membranes are fluid and ionic networks, if existing, are unlikely to develop over long (>5 nm) distances and times. Here we investigate the formation, evolution and impact of such structures on the physical properties of lipid bilayers in solution using single-ion resolution atomic force microscopy (AFM). We find that specific combination of ions alter the diffusivity of lipids in fluid bilayers, and trigger a reversible mesoscale solidification with ~20 nm gel clusters when the membrane is placed under local confinement. The ability of certain ions to induce the formation of these ‘stress clusters’ is inversely proportional to the mobility of the lipids, suggesting ions to create local clustering of the lipids that subsequently behave as a local solid-like diffusing object. While the confinement is here induced by the AFM tip, in biological systems this could be related to contact points with the cytoskeleton or extracellular matrix, or particular proteins.
The impact of these ionic effects on interfacial process could be directly demonstrated on the adsorption of biomolecules to the membrane. Results with antimicrobial peptides and common drugs suggest show correlation between the site of adsorption and the structure of the interface.
3. The interplay between adsorbed ions and membrane curvature
Biomembranes are 3-dimensional objects that are free to fluctuate in solution. Membrane curvature is key to many cellular processes from endocytosis to cell division and protein segregation. Here we examined the effect of curvature by systematically measuring the surface potential (linked to ionic adsorption) of vesicles made with varying diameter. The results consistently show a progressive but dramatic decrease of the vesicles’ surface potential for diameter smaller than ~150 nm, thereby demonstrating that curvature significantly impact ion’s adsorption. The size of the transition curvature show that this is an entropic effect, induced by a stressing of the local hydrogen bond network seems to be at play, an interpretation confirmed by the presence of ionic specificity and a dependence on the type of lipids at play.
These results are significant for medical research because the vesicles are comparable in size to exosomes, which mediate cellular communication. Curvature effects may play an important role in the stability of exosomes and hence provide an important avenue for the development of cancer therapies based on synthetic extracellular vesicles.
4. The impact of electric field on the nanoscale structure of bilayers. Under natural conditions, biomembranes are exposed to a transmembrane electrical potential of typically 40 to 80mV.
Electric fields applied across the membrane affect the behaviour of the bilayer and are able to induce nanoscale structuring of the lipids and changes in the lipids diffusivity depending on the nature of the lipids and the ions involved. Here, fluid membranes were examined in different ionic solutions. The results show a significant impact of both ions (without electric field) and electrical potentials on the behaviour and nanoscale structure of the membranes, but with ions showing the dominating effect. Ion-specific effects are also common, as expected from previous studies.
Overall, we expect the primary impact of this work to be of fundamental nature by challenging and re-shaping our understanding of biointerfaces and the associated energetics. However, the fundamental and universal nature of many of the results suggest that they are transferable to other systems, beyond lipid membranes. Additionally, the uncovered effects can in principle be exploited for technology, for example in the development of environmentally friendly functional interfaces that can prevent bio-fouling, or for informing novel biomedical developments based on lipid structures.
We expect the primary beneficiary of the present project to be other academics and researchers
Contact: kislon.voitchovsky@durham.ac.uk
The project is organised along 4 main research lines:
1. Formation of ionic networks at the surface of gel-phase lipid bilayers.
The existence of nanoscale ionic networks has never been observed at the surface of lipid membranes. Using ‘static’ gel-phase membrane offer an ideal starting point to showing the existence of possible networks and investigate their properties.
Several important results have stemmed from the project. Arguably the main result is the confirmation of postulated long-lived ionic networks at the surface of charged membranes. Atomic-level images in solution show that the networks strongly depend on the types of ions involved both in size and stability. These ionic networks evolve remarkably slowly, typically over seconds to minutes. Computer simulations showed that these mesoscale clusters are ion-specific and their cohesion originates from water-mediated interfaces involving the adsorbed ions in different hydration states and the lipid headgroups. We demonstrate the networks to locally change the stiffness of the membrane by more than 10%, effectively providing a mechanism for membranes to passively modulate their mechanical properties at the nanoscale in a spontaneous manner (without added external energy).
These results are unprecedented and suggest that our current understanding of biological membranes misses an important regulation mechanism. Our current understanding of the regulation of membrane shape, mechanical and viscous properties involve specialized proteins and molecules. This project demonstrates that interfacial effects can be sufficient to achieve similar results, and therefore need to be considered in any comprehensive model of biomembranes. We foresee the results to have a direct impact on both fundamental science and socio-economical applications. First the novelty and groundbreaking nature of our results is expected to lead to several significant scientific publications and presentations in conferences and meeting. The results are also expected to change our current understanding of biological membranes by including interfacial effects as an integral part of the membrane. We expect this view to eventually find its way into teaching manuals and become part of the standard model accepted by students and specialists alike. Second, the results will have implications for the pharmaceutical industry, in particular for the development and testing of new drugs that can exploit the reported ionic networks.
2. Impact of ions on the behaviour of fluid lipid membranes.
In most natural systems, membranes are fluid and ionic networks, if existing, are unlikely to develop over long (>5 nm) distances and times. Here we investigate the formation, evolution and impact of such structures on the physical properties of lipid bilayers in solution using single-ion resolution atomic force microscopy (AFM). We find that specific combination of ions alter the diffusivity of lipids in fluid bilayers, and trigger a reversible mesoscale solidification with ~20 nm gel clusters when the membrane is placed under local confinement. The ability of certain ions to induce the formation of these ‘stress clusters’ is inversely proportional to the mobility of the lipids, suggesting ions to create local clustering of the lipids that subsequently behave as a local solid-like diffusing object. While the confinement is here induced by the AFM tip, in biological systems this could be related to contact points with the cytoskeleton or extracellular matrix, or particular proteins.
The impact of these ionic effects on interfacial process could be directly demonstrated on the adsorption of biomolecules to the membrane. Results with antimicrobial peptides and common drugs suggest show correlation between the site of adsorption and the structure of the interface.
3. The interplay between adsorbed ions and membrane curvature
Biomembranes are 3-dimensional objects that are free to fluctuate in solution. Membrane curvature is key to many cellular processes from endocytosis to cell division and protein segregation. Here we examined the effect of curvature by systematically measuring the surface potential (linked to ionic adsorption) of vesicles made with varying diameter. The results consistently show a progressive but dramatic decrease of the vesicles’ surface potential for diameter smaller than ~150 nm, thereby demonstrating that curvature significantly impact ion’s adsorption. The size of the transition curvature show that this is an entropic effect, induced by a stressing of the local hydrogen bond network seems to be at play, an interpretation confirmed by the presence of ionic specificity and a dependence on the type of lipids at play.
These results are significant for medical research because the vesicles are comparable in size to exosomes, which mediate cellular communication. Curvature effects may play an important role in the stability of exosomes and hence provide an important avenue for the development of cancer therapies based on synthetic extracellular vesicles.
4. The impact of electric field on the nanoscale structure of bilayers. Under natural conditions, biomembranes are exposed to a transmembrane electrical potential of typically 40 to 80mV.
Electric fields applied across the membrane affect the behaviour of the bilayer and are able to induce nanoscale structuring of the lipids and changes in the lipids diffusivity depending on the nature of the lipids and the ions involved. Here, fluid membranes were examined in different ionic solutions. The results show a significant impact of both ions (without electric field) and electrical potentials on the behaviour and nanoscale structure of the membranes, but with ions showing the dominating effect. Ion-specific effects are also common, as expected from previous studies.
Overall, we expect the primary impact of this work to be of fundamental nature by challenging and re-shaping our understanding of biointerfaces and the associated energetics. However, the fundamental and universal nature of many of the results suggest that they are transferable to other systems, beyond lipid membranes. Additionally, the uncovered effects can in principle be exploited for technology, for example in the development of environmentally friendly functional interfaces that can prevent bio-fouling, or for informing novel biomedical developments based on lipid structures.
We expect the primary beneficiary of the present project to be other academics and researchers
Contact: kislon.voitchovsky@durham.ac.uk