Nanoscopic Domains Research Articles

Nanoscopic lipid domains determined by microscopy and neutron scattering

Biological membranes are highly complex supramolecular assemblies, which play central roles in biology. However, their complexity makes them challenging to study their nanoscale structures. To overcome this challenge, model membranes assembled using reduced sets of membrane-associated biomolecules have been found to be both excellent and tractable proxies for biological membranes. Due to their relative simplicity, they have been studied using a range of biophysical characterization techniques. In this review article, we will briefly detail the use of fluorescence and electron microscopies, and X-ray and neutron scattering techniques used over the past few decades to study the nanostructure of biological membranes.

Membrane lipid modulations by methyl-β-cyclodextrin uncouple the Drosophila light-activated phospholipase C from TRP and TRPL channel gating

Sterols are hydrophobic molecules, known to cluster signaling membrane-proteins in lipid-rafts, while methyl-β-cyclodextrin (MβCD) has been a major tool for modulating membrane-sterol content to study its effect on membrane proteins, including the Transient Receptor Potential (TRP) channels. The Drosophila light-sensitive TRP channels are activated downstream of a G-protein-coupled phospholipase Cβ (PLC) cascade. In phototransduction, PLC is a critical enzyme that hydrolyzes phosphatidylinositol 4,5-bisphosphate (PIP2) generating diacylglycerol (DAG), inositol-tris-phosphate (IP3) and protons, leading to TRP and TRP-like (TRPL) channel openings. Here we studied the effects of MβCD on Drosophila phototransduction using electrophysiology while fluorescently-monitoring PIP2 hydrolysis, aiming to examine the effects of sterol modulation on PIP2 hydrolysis and the ensuing light-response in the native system. Incubation of photoreceptor cells with MβCD dramatically reduced the amplitude and kinetics of the TRP/TRPL-mediated light-response. MβCD also suppressed PLC-dependent TRP/TRPL constitutive channel activity in the dark induced by mitochondrial uncouplers, but PLC independent activation of the channels by linoleic acid was not affected. Furthermore, MβCD suppressed a constitutively-active TRP mutant-channel, trpP365, suggesting that TRP channel activity is a target of MβCD action. Importantly, whole-cell voltage-clamp measurements from photoreceptor cells and simultaneously monitored PIP2 hydrolysis by translocation of fluorescently-tagged lipid-binding-Tubby-protein domain, from the plasma membrane to the cytosol revealed that MβCD virtually abolished the light-response when having only little effect on the light-activated PIP2 hydrolysis by PLC. Together, MβCD uncoupled TRP/TRPL channel gating from light-activated PLC and PIP2-hydrolysis suggest the involvement of distinct nanoscopic lipid domains such as lipid-rafts and PIP2-clusters in TRP/TRPL channel gating.

On the force between "rafts".

"Rafts" in the plasma membrane are nanoscopic domains rich in sphingolipids and cholesterol. Groups of them are thought to provide stable platforms for various proteins. How several rafts can form a larger stable platform has yet to be made clear. Here, we show that in the microemulsion theory of rafts, in which they, and the sea in which they float, are characterized by a naturally occurring length, rafts separated by a sea larger than this length will attract one another, thus making the sea smaller. Similarly, if rafts are separated by a sea that is smaller than the characteristic size, they will repel one another thus allowing the sea to increase in size. Such an elastic force can clearly stabilize a platform of many rafts.

Evidence for ferroelastic switching and nanoscopic domains in polycrystalline Si-doped hafnium oxide films

The mechanism of nanoscopic domain switching in ferroelectric hafnium oxide and its implications for antiferroelectric-like behavior as well as for the wake-up effect is still widely discussed. Understanding this mechanism is of vital importance for a multitude of applications like piezoelectric actuators, pyroelectric sensors, and nonvolatile memory devices. In this article, electrical and physical analysis methods are used to characterize ferroelectric hafnium oxide on the nanoscopic as well as the macroscopic length scale. Evidence for nanoscopic domains is found using transmission Kikuchi diffraction. In combination with macroscopic Preisach density measurements, strong evidence is found that antiferroelectric-like behavior and wake-up are governed by ferroelastic switching, i.e., a 90° domain wall motion. Based on these insights, the material stack can be optimized to further improve microelectronic applications based on HfO2.

Which Moiety Drives Gangliosides to Form Nanodomains?

Gangliosides are important glycosphingolipids involved in a multitude of physiological functions. From a physicochemical standpoint, this is related to their ability to self-organize into nanoscopic domains, even at molar concentrations of one per 1000 lipid molecules. Despite recent experimental and theoretical efforts suggesting that a hydrogen bonding network is crucial for nanodomain stability, the specific ganglioside moiety decisive for the development of these nanodomains has not yet been identified. Here, we combine an experimental technique achieving nanometer resolution (Förster resonance energy transfer analyzed by Monte Carlo simulations) with atomistic molecular dynamic simulations to demonstrate that the sialic acid (Sia) residue(s) at the oligosaccharide headgroup dominates the hydrogen bonding network between gangliosides, driving the formation of nanodomains even in the absence of cholesterol or sphingomyelin. Consequently, the clustering pattern of asialoGM1, a Sia-depleted glycosphingolipid bearing three glyco moieties, is more similar to that of structurally distant sphingomyelin than that of the closely related gangliosides GM1 and GD1a with one and two Sia groups, respectively.

Messenger Materials Moving Forward: The Role of Functional Polymer Architectures as Enablers for Dynamic Nano‐to‐Macro Messaging

AbstractIdentifying changes in the nanoscopic domain is a key challenge in the physicochemical sciences, where great interest is on sensing complex processes that involve cellular biochemical reactions, chemical heterogeneities, contact forces, and other interfacial phenomena. This has stimulated the development of diverse materials that allow subtle nanoscopic environments to be "seen". The challenge in the nano‐domain has always been the ability to sense changes on the minute scale and rapidly transduce the information out for macroscopical observation. Ideally, materials should inform when processes are occurring. Recently, new systems that leverage established concepts with fluorescence‐ and plasmonic‐based sensing have been devised, which has reinvigorated the domain, where functional polymers coupled in specific architectures to transducing motifs allow for a new basis of messenger materials to be realized. The key aspect in this regard is that the polymers allow for sensing to be achieved only when they are carefully coupled to the amplification system. In this perspective, the role of specific functional polymer architectures for the realization of nano‐to‐macro sensing of subtle nano‐messengers is discussed and where the exciting field of messenger materials is seen moving forward is pointed out.

Repairable Macroscopic Monodomain Arrays from Block Copolymers Enabled by Photoplastic and Photodielectric Effects

Upon exposure to UV light (120 mW/cm2, λ = 365 nm), a trans-cis isomerization occurs in a cylinder-forming, azobenzene-containing block copolymer of polydimethylsiloxane-b-poly((4(phenyldiazenyl)phenoxy)hexyl acrylate) (PDMS-b-PPHA) that enables the generation of monodomains of healable, long-range ordered arrays of nanoscopic domains over macroscopic distances. The trans-cis isomerization gives rise to a significant increase in the dielectric constant (from 6.52 to 19.8 at 100 Hz, photodielectric behavior) and a dramatic decrease in the Tg (from 54 to 1 °C, photoplastic behavior) of the PPHA block. By combining these characteristics with an in-plane electric field, macroscopic monodomains of near-perfectly aligned cylindrical microdomains are achieved at low temperatures, and a damage repair is clearly uncovered, where the 300 nm wide scratches can be completely healed at 40 °C, leaving a smooth, uniformly thick film where the continuity and orientation of the aligned microdomains are restored. Subsequent exposure to visible light causes a cis-trans isomerization, increasing the matrix Tg to 54 °C, producing highly oriented and aligned PDMS cylindrical microdomains in a PPHA matrix.

Cholesterol Regulation of Membrane Proteins Revealed by Two-Color Super-Resolution Imaging.

Cholesterol and phosphatidyl inositol 4,5-bisphosphate (PIP2) are hydrophobic molecules that regulate protein function in the plasma membrane of all cells. In this review, we discuss how changes in cholesterol concentration cause nanoscopic (<200 nm) movements of membrane proteins to regulate their function. Cholesterol is known to cluster many membrane proteins (often palmitoylated proteins) with long-chain saturated lipids. Although PIP2 is better known for gating ion channels, in this review, we will discuss a second independent function as a regulator of nanoscopic protein movement that opposes cholesterol clustering. The understanding of the movement of proteins between nanoscopic lipid domains emerged largely through the recent advent of super-resolution imaging and the establishment of two-color techniques to label lipids separate from proteins. We discuss the labeling techniques for imaging, their strengths and weakness, and how they are used to reveal novel mechanisms for an ion channel, transporter, and enzyme function. Among the mechanisms, we describe substrate and ligand presentation and their ability to activate enzymes, gate channels, and transporters rapidly and potently. Finally, we define cholesterol-regulated proteins (CRP) and discuss the role of PIP2 in opposing the regulation of cholesterol, as seen through super-resolution imaging.

3D Printed Solid Polymer Electrolytes with Bicontinuous Nanoscopic Domains for Ionic Liquid Conduction and Energy Storage.

Solid polymer electrolytes (SPEs) offer several advantages compared to their liquid counterparts, and much research has focused on developing SPEs with enhanced mechanical properties while maintaining high ionic conductivities. The recently developed polymerization-induced microphase separation (PIMS) technique offers a straightforward pathway to fabricate bicontinuous nanostructured materials in which the mechanical properties and conductivity can be independently tuned. In this work SPEs with tunable mechanical properties and conductivities are prepared via digital light processing 3D printing, exploiting the PIMS process to achieve nanostructured ion-conducting materials for energy storage applications. A rigid crosslinked poly(isobornyl acrylate-stat-trimethylpropane triacrylate) scaffold provided materials with room temperature shear modulus above 400MPa, while soft poly(oligoethylene glycol methyl ether acrylate) domains containing the ionic liquid 1-butyl-3-methylimidazolium bis-(trifluoromethyl sulfonyl)imide endowed the material with ionic conductivity up to 1.2 mS cm-1 at 30°C. These features make the 3D-printed SPE very competitive for applications in all solid energy storage devices, including supercapacitors.

The use of iodinated cholesterol to enhance contrast in cryo-EM images of lipid membranes.

The lipid raft hypothesis posits nanoscopic domains in the plasma membrane (PM) that arise from unfavorable interactions between low-melting phospholipids and high-melting sphingomyelins in the PM outer leaflet. Cholesterol modulates the properties of these domains (for example, ensuring their fluidity) and can influence the coupling of phase behavior between the outer and inner leaflets. However, the interleaflet distribution of cholesterol has proven difficult to measure. Cryogenic electron microscopy (cryo-EM) is among the few techniques capable of directly visualizing the cholesterol distribution, but this would require an electron-dense cholesterol analog that faithfully mimics the properties of native cholesterol. Here, we investigated a variant of cholesterol in which one hydrogen on the C19 methyl group was replaced with iodine. As expected, cryo-EM images of model membranes containing cholesterol-I19 revealed enhanced contrast compared to vesicles with native cholesterol. However, fluorescence microscopy of GUVs showed that cholesterol-I19 does not support macroscopic liquid-ordered+liquid-disordered phase separation in ternary mixtures with DPPC and DOPC, although FRET and cryo-EM measurements were consistent with nanoscopic heterogeneity in compositions that appeared uniform in GUVs.

Optimization of cryo-electron microscopy for quantitative analysis of lipid bilayers

Cryogenic electron microscopy (cryo-EM) is among the most powerful tools available for interrogating nanoscale structure of biological materials. We recently showed that cryo-EM can be used to measure the bilayer thickness of lipid vesicles and biological membranes with subangstrom precision, resulting in the direct visualization of nanoscopic domains of different thickness in multicomponent lipid mixtures and giant plasma membrane vesicles. Despite the great potential of cryo-EM for revealing the lateral organization of biomembranes, a large parameter space of experimental conditions remains to be optimized. Here, we systematically investigate the influence of instrument parameters and image postprocessing steps on the ability to accurately measure bilayer thickness and discriminate regions of different thickness within unilamellar liposomes. This unique application of cryo-EM places particular demands on image acquisition optimization and analysis due to the facts that 1) each vesicle is a different size with different curvature, 2) the domains in each vesicle can be heterogenous in size, and 3) the random orientation of vesicles amplifies the variability of domain size in projected images. We also demonstrate a spatial autocorrelation analysis to extract additional information about lateral heterogeneity.

Patterning of large area nanoscale domains in as-grown epitaxial ferroelectric PbTiO3 films

Effective tuning of nanoscale domain structures provides fundamental basis for controlling and engineering of various functionalities in ferroelectric materials. In this work, we demonstrate the precise patterning of nanoscopic domain structures in as-grown epitaxial PbTiO3 (PTO) films by merely introducing an ultrathin pre-patterned doping layer (e.g., Fe-doped PTO). The doping layer can effectively reverse the interfacial built-in bias, consequent to a reversed initial polarization reorientation in the as-grown film, which makes it possible to transfer the nano-patterns in the doping layer into the domain structure of ferroelectric films. For instance, we have successfully fabricated large area ordered array of nanoscale cylindrical domains (downward polarization) embedded in the matrix domain with opposite polarization (upward polarization) in PTO film. These nanoscale cylinder domains also allow deterministic and reversible erasure and creation induced by biased tip scanning. The results provide an effective pathway for on-demand patterning of large area nanoscale domains in the as-grown films, which may find applications in a wide range of nanoelectronic devices.

Spontaneously formed multiscale nano-domains in monophasic region of ternary solution

HypothesisTernary mixtures, containing one hydrotrope and two immiscible fluids, both being soluble in the hydrotrope at any proportion, exhibit unexpected solubilization power and unusual mesoscopic properties, which are a subject of long-standing controversies for decades. ExperimentsThis work investigates the entire monophasic region of ternary trans-anethol/ethanol/water system, where multiscale nanostructurings with correlation lengths exceeding dimensions of individual molecules are identified by dynamic light scattering. The physical properties of the ternary mixture are characterized, with revealing the compositional dependence of refractive index and dynamic viscosity. FindingsIn surfactant-free microemulsion (SFME) regime, the single phase consists of two distinct nanoscopic domains in equilibrium, one trans-anethol-rich aggregate at the molecular scale (∼1 nm) and one mesoscopic droplet at the mesoscale (∼100 nm). However, only a tiny fraction of the hydrophobic component trans-anethol (<∼0.025%) are initially incorporated in mesoscale structures, and the vast majority are molecularly dissolved or in the form of aggregates. The concentration of mesoscopic droplets increases sharply when shifting toward the miscibility gap, but their size exhibits a weak compositional dependence. The nanoscopic domains exhibit long-term physical stability, and the Ostwald ripening is still the primary mechanism governing such mesoscopic droplet aging, but with a rather low ripening rate.

Sodiation-Induced Electrochromism in Carbon Nanofoam–Paper Electrodes

Carbon nanofoam papers (CNFPs) serve as device-ready negative electrodes for nonaqueous sodium-ion (Na-ion) batteries, with fast and reversible Na+ storage at the aerogel-like, disordered carbon nanofoam. These binder-free, scalable electrode architectures are not only advantageous for the construction of practical, high-performance Na-ion cells, but are also effective platforms to analyze charge-storage mechanisms. We use in situ optical imaging at the outer surfaces of CNFPs in a Na half-cell to track the distinctive color changes that accompany the multi-stage Na+-storage processes on scanning these electrodes through their active voltage window (0–2 V vs Na∣Na+). We observe that electrochromic transitions ― black in the native (unsodiated) state→blue→red/gold ― occur primarily with deep levels of sodiation at <0.5 V vs Na∣Na+. In situ Raman scattering measurements in the same optical cell show that these color changes correlate with shifts in the characteristic G-band Raman peak that would indicate bulk Na+ insertion into nanoscopic graphitic domains within the aerogel-like carbon. The CNFPs also exhibit appreciable Na-ion storage at higher voltage (0.5–2 V vs Na∣Na+), which can be ascribed to surface-based mechanisms that are accompanied by shifts in the D-band Raman peak, but which do not induce appreciable color change.

Influence of ceramide on lipid domain stability studied with small-angle neutron scattering: The role of acyl chain length and unsaturation

Ceramides and diacylglycerols are groups of lipids capable of nucleating and stabilizing ordered lipid domains, structures that have been implicated in a range of biological processes. Previous studies have used fluorescence reporter molecules to explore the influence of ceramide acyl chain structure on sphingolipid-rich ordered phases. Here, we use small-angle neutron scattering (SANS) to examine the ability of ceramides and diacylglycerols to promote lipid domain formation in the well-characterized domain-forming mixture DPPC/DOPC/cholesterol. SANS is a powerful, probe-free technique for interrogating membrane heterogeneity, as it is differentially sensitive to hydrogen’s stable isotopes protium and deuterium. Specifically, neutron contrast is generated through selective deuteration of lipid species, thus enabling the detection of nanoscopic domains enriched in deuterated saturated lipids dispersed in a matrix of protiated unsaturated lipids. Using large unilamellar vesicles, we found that upon replacing 10 mol% DPPC with either C16:0 or C18:0 ceramide, or 16:0 diacylglycerol (dag), lipid domains persisted to higher temperatures. However, when DPPC was replaced with short chain (C6:0 or C12:0) or very long chain (C24:0) ceramides, or ceramides with unsaturated acyl chains of any length (C6:1(3), C6:1(5), C18:1, and C24:1), as well as C18:1-dag, lipid domains were destabilized, melting at lower temperatures than those in the DPPC/DOPC/cholesterol system. These results show how ceramide acyl chain length and unsaturation influence lipid domains and have implications for how cell membranes might modify their function through the generation of different ceramide species.

Evidence that cuprate superconductors form an array of nanoscopic Josephson junctions

Recent measurements of charge instabilities in overdoped compounds rekindled the proposal that cuprates become superconductors by long-range order through Josephson coupling between nanoscopic charge domains. We use the theory of phase-ordering dynamics to show that incommensurate charge density waves (CDWs) are formed in the CuO planes by a series of free-energy wells separated by steep barriers. Charge oscillations in these domains give rise to a net hole-hole attraction proportional to the height of these barriers. Concomitantly, the self-consistent calculations yield localized superconducting amplitudes in the CDW domains characterizing a granular superconductor. We show that a transition by long-range phase order promoted by Josephson coupling elucidates many well-known features of cuprates like the high magnetic penetration depth anisotropy and the origin of the pseudogap, among others. Furthermore, the average Josephson energy reproduces closely the planar superfluid density temperature dependence of La-based films and the superconducting giant proximity effects of cuprates, a 20-year-old open problem.

Environmental Exploration of Ultra-Dense Nanobubbles: Rethinking Sustainability

Nanobubbles are nanoscopic gaseous domains than can exist on solid surfaces or in bulk liquids. They have attracted significant attention in the last decade due to their long-time (meta)stability and ready potential for real-world applications, especially in environmental engineering and more sustainable ecosystems, water treatment, irrigation, and crop growth. After reviewing important nano-bubble science and activity, with some of the latest promising results in agriculture, we point out important directions in applications of nano-bubble phenomena for boosting sustainability, with viewpoints on how to revolutionise best-practice environmental and green sustainability, taking into account economic drivers and impacts. More specifically, it is pointed out how nanobubbles may be used as delivery vehicles, or “nano-carriers”, for nutrients or other agents to specific targets in a variety of ecosystems of environmental relevance, and how core this is to realising a vision of ultra-dense NBs in shaping a positive and lasting impact on ecosystems and our natural environment.

Insights of phase separation in bilayers of ternary mixtures with different sterols

It has been shown that polyene antimycotics are selective according to the sterol present in the membrane, leading to stronger activity in fungal cells. However, it is not clear the role of the membrane structure in ionophoretic activity. In recent years, there has been a debate about if the homogenous model of the membrane is accurate or there exist nanoscopic domains enriched with sphingomyelin and cholesterol. The influence of sterol and its interaction with sphingomyelin can provide information on ordered domain formation.

Remotely controllable supramolecular rotor mounted inside a porphyrinic cage

•Design and synthesis of a supramolecular rotor within a porous Zn–porphyrinic cage•High-yielding IEDDA reaction enables facile construction of the rotor inside the cage•Initiation of both rotary and tumbling motions of the rotor with chemical stimuli•Reversible control of the dual mechanical motions by a photo-dissociable ligand The confinement of biological molecular machines, which are controlled by various chemical or physical stimuli within the cytoskeleton matrix, endows them with remarkable precision and programmable mechanical motions. Biological molecular machines are far more complex than any artificial molecular machine ever built. To mimic the intricate behavior of biological machinery, strategies need to be developed for embedding and confining artificial molecular machines within nanoscopic domains in which their functions can be specifically controlled by external stimuli. One such approach is the confinement of molecular machines inside porous organic cages. The high porosity of the organic cages provides ample room for uninterrupted movement, and its shape-persistent and interactive framework provides a high degree of control over the mechanical motion triggered by external stimuli. Our strategy may lay the foundation for the development of tunable molecular devices using porous organic cages. The confinement of molecular machines into nanostructured cages and controlling their functions by external stimuli holds great potential for the creation of smart functional materials that imitate the embodied intelligence of biological processes. Herein, we report the construction of a supramolecular rotor in a porous Zn-metallated porphyrinic cage (1) by encapsulation of a tetrazine-based linear axle (LA) via metal-ligand coordination bond, followed by post-assembly modification to append a controllable side arm to LA via inverse electron demand Diels-Alder (IEDDA) reaction. While the rotor alone shows nearly no motion, the addition of pyridine derivatives as a zinc coordinating ligand results in both 90° jump-like rotary motion of the rotor and slow tumbling motion of the rotor axle in a stochastic manner. Interestingly, the dual motions of the rotor can be reversibly controlled by the UV and visible light-induced coordination and dissociation of an azopyridine-based ligand with Zn centers as a signal transducer. The confinement of molecular machines into nanostructured cages and controlling their functions by external stimuli holds great potential for the creation of smart functional materials that imitate the embodied intelligence of biological processes. 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Berrocal J.A. Vanthuyne N. Rutjes F.P.J.T. Feringa B.L. Elemans J.A.A.W. Nolte R.J.M. Molecular motor-functionalized porphyrin macrocycles.Nat. Commun. 2020; 11: 5291Google Scholar Despite these inherent advantages, it still remains synthetically challenging to install molecular machines inside the nano-sized porous organic cages and to control the molecular motion by chemical or/and physical stimuli. Of particular interest is controlling confined molecular machines remotely by an external stimulus, which may unlock new properties of these systems with spatiotemporal precision. In comparison with other remotely controlled stimuli, light has unique advantages of being a clean energy source and having the ability to be transiently delivered at a precise location in a solution.38van Leeuwen T. Lubbe A.S. Štacko P. Wezenberg S.J. Feringa B.L. Dynamic control of function by light-driven molecular motors.Nat. Chem. Rev. 2017; 1: 96Google Scholar,39Kundu P.K. Samanta D. Leizrowice R. 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However, using a light source to control a molecular machine that is devoid of any photoresponsive component can be very interesting considering its resemblance with the biological machines, e.g., light-driven ion pumps. We have recently reported cube-shaped porous organic cages self-assembled from six square-shaped porphyrins and eight triangular linkers by dynamic covalent chemistry.52Mukhopadhyay R.D. Kim Y. Koo J. Kim K. Porphyrin boxes.Acc. Chem. Res. 2018; 51: 2730-2738Google Scholar, 53Benke B.P. Aich P. Kim Y. Kim K.L. Rohman M.R. Hong S. Hwang I.-C. Lee E.H. Roh J.H. Kim K. Iodide-selective synthetic ion channels based on shape-persistent organic cages.J. Am. Chem. Soc. 2017; 139: 7432-7435Google Scholar, 54Hong S. Rohman M.R. Jia J. Kim Y. Moon D. Kim Y. Ko Y.H. Lee E. Kim K. Porphyrin boxes: rationally designed porous organic cages.Angew. Chem. Int. Ed. Engl. 2015; 54: 13241-13244Google Scholar Shape-persistent porphyrinic cages, due to their well-defined receptor cavities, are ideal platforms to install molecular machines in a precise and predictable manner. Previously, a number of Zn–porphyrins-based dynamic molecular machines have been reported.55Goswami A. Schmittel M. Double rotors with fluxional axles: domino rotation and azide–alkyne Huisgen cycloaddition catalysis.Angew. Chem. Int. Ed. Engl. 2020; 59: 12362-12366Google Scholar, 56Gaikwad S. Goswami A. De S. Schmittel M. A Metalloregulated four-state nanoswitch controls two-step sequential catalysis in an eleven-component system.Angew. Chem. Int. Ed. Engl. 2016; 55: 10512-10517Google Scholar, 57Liu S. Kondratuk D.V. Rousseaux S.A.L. Gil-Ramírez G. O'Sullivan M.C. Cremers J. Claridge T.D.W. Anderson H.L. Caterpillar track complexes in template-directed synthesis and correlated molecular motion.Angew. Chem. Int. Ed. Engl. 2015; 54: 5355-5359Google Scholar, 58Muraoka T. Kinbara K. Aida T. Mechanical twisting of a guest by a photoresponsive host.Nature. 2006; 440: 512-515Google Scholar However, the establishment of a rational and high-yielding synthetic strategy to precisely organize molecular machines inside Zn–porphyrin-based porous organic cages with a 3D encompassed intrinsic nanoscopic cavity and to control their functions with multiple external stimuli requires further exploration. Herein, we report the synthesis of a supramolecular rotor built inside a porous Zn–porphyrinic cage (1) by insertion of a tetrazine-based LA followed by post-assembly modification to append a controllable side arm to the axle. The 1H and 2D EXSY NMR studies reveal a 90° jump-like random rotary motion of the rotor arm and a slow tumbling motion of the rotor axle upon the addition of pyridine derivatives, which compete with the rotor for zinc–porphyrin coordination. Computational calculations confirm the stochastic nature of the molecular motions. Moreover, the rotation rates of these two motions can be adjusted by varying the amounts of pyridine derivatives. The reversible control over the dual motions of the rotor is achieved by UV and visible light-induced coordination and dissociation of azopyridine-based photo-dissociable ligand (PDL), which acts as a signal transducer. The nanoscale hydrophobic cavity of the Zn–porphyrinic cage and the remotely controllable functions of the rotor may collectively lead to the development of tunable molecular devices with potential applications in reversible on-off ion channels, switchable catalysis, and drug delivery. At the outset of this work, we synthesized a pyridine terminated LA (length, ∼1.9 nm, see synthesis in Scheme S1), which can coordinate to two Zn metal centers of 1 (distance, ∼2.3 nm), across the cavity (Figure S1). LA consists of a central tetrazine moiety that can react efficiently with alkenes or alkynes dienophiles via inverse electron demand Diels-Alder (IEDDA) reaction to attach a new functionality on LA. By using the remarkably high-yielding IEDDA reactions for post-assembly modification, a number of different rotor moieties can be precisely incorporated into the cage. We envisaged that post-assembly modification of LA inside 1 with (1R,8S,9s)-bicyclo[6.1.0]-non-4-yn-9-ylmethanol (L1) may provide a controllable side arm for the freely rotating axle as L1 contains a hydroxy group (–OH) at the terminal position, which can coordinate with the “equatorial” Zn–porphyrins of 1 (Figure 1). The mechanical motion of the resulting assembly L1-LA⊂1 can be triggered in the presence of chemical stimuli, such as pyridine derivatives, which can coordinate with the Zn–porphyrinic faces of L1-LA⊂1 from outside the hydrophobic cavity, thus initiating the motion by inhibiting the formation of coordination bonds between the Zn centers of 1 and the rotor L1-LA (Figure 1B). The axle insertion experiment was performed by adding a stoichiometric amount of LA to the chloroform solution of 1, yielding LA⊂1 (Figure 1A). The binding affinity (Ka) of 1 for LA was determined to be 3.8 ± 0.1 × 107 M−1 by UV-vis titration (Figure S2), confirming a strong host-guest interaction between 1 and LA. In the 1H NMR spectrum of LA⊂1 in toluene-d8, LA protons showed a large upfield shift compared with their free state, due to the strong ring current of the porphyrin units. In particular, the α-py protons that are closest to the porphyrin ring showed the largest upfield shift (8.84 to 1.88 ppm), indicating that LA is precisely positioned inside 1 (Figure S3). Moreover, the insertion of LA into the cavity of 1 splits the identical pyrrolic protons (a) of all six porphyrins of 1 into three sets: a1 protons for two “axial” porphyrins (connecting to the axle) and a2 and a3 protons for four “equatorial” porphyrins (parallel to the axle). The a2 and a3 protons show different chemical shifts as they are directed toward the axial and equatorial porphyrins, respectively (Figure 2). The NMR splitting pattern of the cage protons suggests the O → D4 desymmetrization of the cage structure. All peaks were assigned by several 2D NMR experiments (1H–13C HSQC [heteronuclear single quantum coherence spectroscopy], 1H–1H COSY [homonuclear correlation spectroscopy], and ROESY [rotating-frame nuclear overhauser effect spectroscopy]; Figures S4–S7). A single trace in the DOSY (diffusion-ordered spectroscopy) spectrum confirmed the presence of a single species (LA⊂1) in the solution (Figure S8). Finally, X-ray structure analysis of LA⊂1 (for crystallization conditions, see supplemental information) confirmed the coordination of the py moieties of LA to the Zn metal centers of the opposite faces of 1 (Zn–Npy = 2.081(5) Å). In the crystal structure, LA⊂1 maintained an overall cube-shape with the same opposite Zn⋅⋅⋅Zn distance (23.8 Å) in all three directions despite the insertion of LA (Figure 2A). To demonstrate facile post-assembly modification of LA⊂1, norbornadiene was chosen as a model compound for IEDDA reaction that quantitatively converts the tetrazine-based axle (LA) to a pyridazine-based axle (PLA) inside 1 (PLA⊂1) within 4 h at room temperature, which is manifested by the disappearance of 1H NMR signals of LA and the appearance of PLA signals (Figures S9 and S10). The formation of PLA⊂1 was confirmed by 2D NMR and DOSY analysis (Figures S11–S13). The post-assembly modification of LA⊂1 with L1 substrate (1 equiv per tetrazine) by IEDDA reaction proceeded almost instantly at room temperature to furnish a ring-fused pyridazine-based axle inside 1 (L1-LA⊂1), as verified by 1H NMR, MALDI-TOF, and elemental analysis (EA) (Figures S14 and S15). Thermal stability was confirmed by thermogravimetric analysis (TGA) (Figures S16 and S17). In the 1H NMR spectrum of L1-LA⊂1 in toluene-d8, the L1 protons are shifted in the upfield region (appeared between 0 to −5 ppm), which may be due to their close proximity with one of the “equatorial” Zn–porphyrin from inside the cavity. In addition, the pyrrolic protons a3 of the equatorial Zn–porphyrins of LA⊂1 are divided into two sets a3a and a3b, indicating further desymmetrization of the cage structure (Figure 2C). The splitting of phenyl and imine protons also suggests the lowering of cage symmetry. All peaks were assigned by several 2D NMR experiments (Figures S18–S21). Moreover, DOSY experiment confirmed the single species in solution (Figure S22). Unexpectedly, the pyrrolic protons a1 and a2 of L1-LA⊂1 showed no splitting, the reason for which is currently not fully understood. On the other hand, upon directly mixing the 1:1 ratio of L1-LA and 1, a non-symmetrical pattern of LA protons was observed in the 1H NMR spectrum, indicating that L1-LA coordinates to 1 from outside the cage in a monodentate fashion (Figure S23), probably due to small-sized windows of the cage and long alkyl chains blocking the cage windows. After numerous attempts, the structure of L1-LA⊂1 has been finally confirmed by single-crystal X-ray crystallography (Figure 2B). The molecular structure revealed that L1-LA resides inside 1 by the coordination of the py groups of L1-LA to the “axial” Zn–porphyrins (Zn–Npy = 2.129(5) Å) and the hydroxy group (–OH) to the “equatorial” Zn–porphyrin (Zn–OH = 2.379(5) Å). As shown in Figure 2B, the slightly bent axle LA (bent angle, ∼167°) and curved shape of the side arm L1 assist in the coordination of the hydroxy group (–OH) with one of the “equatorial” Zn–porphyrins. The opposite Zn⋅⋅⋅Zn distances in the a and b axes are both 23.7 Å, whereas that in the c axis is 20.1 Å. The shorter Zn⋅⋅⋅Zn distance may be ascribed to the coordination of the –OH group of L1-LA to the equatorial Zn–porphyrin. The bent shape of LA and the curved shape of L1 were also observed in the X-ray crystal structure of L1-LA (Figure S24). Next, we investigated the operation of the supramolecular rotor L1-LA⊂1. In the absence of external stimulus, upon cooling the solution of L1-LA⊂1 (in toluene-d8) down to 243 K, no splitting of pyrrolic protons peaks was observed in the 1H NMR spectrum (Figure S25), which suggests that L1-LA is either rotating too slowly on the NMR timescale or no rotation at all, possibly due to the Zn–OHL1-LA coordination bond. It is known that Zn–porphyrins bind more strongly to pyridine than to alcohols.59Olsson S. Dahlstrand C. Gogoll A. Design of oxophilic metalloporphyrins: an experimental and DFT study of methanol binding.Dalton Trans. 2018; 47: 11572-11585Google Scholar In fact, coordination of pyridine to the Zn center can replace the Zn–OH coordination bond for maintaining a five-coordinate binding mode. The coordination of pyridine to L1-LA⊂1 is borne out by the upfield shift of the pyridine protons peaks in the 1H NMR spectrum. However, in the presence of excess pyridine, the peaks shifted back to the downfield region due to the establishment of a fast equilibrium between coordinated and free pyridine molecules (Figure S26). Thereafter, the effect of the addition of the chemical stimulant (pyridine) to L1-LA⊂1 was studied in detail by 1H NMR spectroscopy. Upon stepwise addition of pyridine (up to 6 equiv) to L1-LA⊂1, the pyrrolic protons a3a and a3b of the four equatorial porphyrins merged into a sharp singlet at 8.98 ppm (Figure 3A). The imine protons and the phenyl protons that correspond to the equatorial porphyrins also merged, indicating the symmetrization of the cage framework (Figure S26A). Additionally, the side arm (L1) protons that appear between 0 to −5 ppm due to their close proximity with one of the equatorial Zn–porphyrins broadened and shifted slightly downfield (Figure S26B). The symmetrical structure of the cage framework as derived from the 1H NMR spectroscopic studies and broadened resonances of the side arm protons suggested that the coordination of pyridine molecules to the Zn centers dissociated the Zn–OH coordination bond, and, as a result, the side arm started jumping rapidly around all four equatorial Zn–porphyrin stations at room temperature by using the rotor axle as a hinge (Figure 1B). To confirm that the mixing of the peaks is due to the rotary motion and not to pyridine coordination, 1H NMR titration between pyridine (0–6 equiv) and 1 was performed, which showed that the coordination of pyridine does not affect the protons signals of cage 1 due to its relatively rapid association/dissociation equilibrium with the Zn–porphyrins of 1 on the NMR timescale (Figure S27). Further insight into the rotor dynamics was obtained from variable-temperature (VT) 1H NMR studies. While the a3a and a3b protons of L1-LA⊂1 appeared as a single peak in the presence of 6 equiv of pyridine at 298 K, the peak was broadened at 263 K and emerged as two distinct peaks at 9.05 and 9.09 ppm at 243 K (Figure 3B). The 1H NMR could not be recorded below 243 K due to poor solubility of L1-LA⊂1 in toluene-d8 at the low temperatures. Upon raising the temperature back to 298 K, the peaks again merged into a single peak at 8.99 ppm (Figure S28). These experiments further corroborated the notion that the rotor exhibits a fast rotary motion in the presence of the chemical stimulant pyridine, whereas the rotation decelerates at low temperatures. Line shape analysis of the VT 1H NMR data in the presence of 6 equi

Structure and dynamics of nanoconfined water and aqueous solutions.

This review is devoted to discussing recent progress on the structure, thermodynamic, reactivity, and dynamics of water and aqueous systems confined within different types of nanopores, synthetic and biological. Currently, this is a branch of water science that has attracted enormous attention of researchers from different fields interested to extend the understanding of the anomalous properties of bulk water to the nanoscopic domain. From a fundamental perspective, the interactions of water and solutes with a confining surface dramatically modify the liquid's structure and, consequently, both its thermodynamical and dynamical behaviors, breaking the validity of the classical thermodynamic and phenomenological description of the transport properties of aqueous systems. Additionally, man-made nanopores and porous materials have emerged as promising solutions to challenging problems such as water purification, biosensing, nanofluidic logic and gating, and energy storage and conversion, while aquaporin, ion channels, and nuclear pore complex nanopores regulate many biological functions such as the conduction of water, the generation of action potentials, and the storage of genetic material. In this work, the more recent experimental and molecular simulations advances in this exciting and rapidly evolving field will be reported and critically discussed.