Geometric Transformations Afforded by Rotational Freedom in Aramid Amphiphile Nanostructures.

Molecular self-assembly in water leads to nanostructure geometries that can be tuned owing to the highly dynamic nature of amphiphiles. There is growing interest in strongly interacting amphiphiles with suppressed dynamics, as they exhibit ultrastability in extreme environments. However, such amphiphiles tend to assume a limited range of geometries upon self-assembly due to the specific spatial packing induced by their strong intermolecular interactions. To overcome this limitation while maintaining structural robustness, we incorporate rotational freedom into the aramid amphiphile molecular design by introducing a diacetylene moiety between two aramid units, resulting in diacetylene aramid amphiphiles (D-AAs). This design strategy enables rotations along the carbon-carbon sp hybridized bonds of an otherwise fixed aramid domain. We show that varying concentrations and equilibration temperatures of D-AA in water lead to self-assembly into four different nanoribbon geometries: short, extended, helical, and twisted nanoribbons, all while maintaining robust structure with thermodynamic stability. We use advanced microscopy, X-ray scattering, spectroscopic techniques, and two-dimensional (2D) NMR to understand the relationship between conformational freedom within strongly interacting amphiphiles and their self-assembly pathways.

Domain-Specific Phase Transitions in a Supramolecular Nanostructure.

Understanding thermal phase behavior within nanomaterials can inform their rational design for medical technologies like drug delivery systems and vaccines, as well as for energy technologies and catalysis. This study resolves thermal phases of discrete domains within a supramolecular aramid amphiphile (AA) nanoribbon. Dynamics are characterized by X-band EPR spectroscopy of spin labels positioned at specific sites through the nanoribbon cross-section. The fitting of the electron paramagnetic resonance (EPR) line shapes reveals distinct conformational dynamics, with fastest dynamics at the surface water layer, intermediate dynamics within the flexible cationic head group domain, and slowest dynamics in the interior aramid domain. Measurement of conformational mobility as a function of temperature reveals first- and second-order phase transitions, with melting transitions observed in the surface and head group domains and a temperature-insensitive crystalline phase in the aramid domain. Arrhenius analysis yields activation energies of diffusion at each site. This work demonstrates that distinct thermal phase behaviors between adjacent nanodomains within a supramolecular nanostructure may be resolved and illustrates the utility of EPR spectroscopy for thermal phase characterization of nanostructures.

Morphological Transitions of a Photoswitchable Aramid Amphiphile Nanostructure.

Self-assembly of small amphiphilic molecules in water can lead to nanostructures of varying geometries with pristine internal molecular organization. Here we introduce a photoswitchable aramid amphiphile (AA), designed to exhibit extensive hydrogen bonding and robust mechanical properties upon self-assembly, while containing a vinylnitrile group for photoinduced cis-trans isomerization. We demonstrate spontaneous self-assembly of the vinylnitrile-containing AA in water to form nanoribbons. Upon UV irradiation, trans-to-cis isomerizations occur concomitantly with a morphological transition from nanoribbons to nanotubes. The nanotube structure persists in water for over six months, stabilized by strong and collective intermolecular interactions. We demonstrate that the nanoribbon-to-nanotube transition is reversible upon heating and that switching between states can be achieved repeatedly. Finally, we use electron microscopy to capture the transition and propose mechanisms for nanoribbon-to-nanotube rearrangement and vice versa. The stability and switchability of photoresponsive AA nanostructures make them viable for a range of future applications.

Dynamics in supramolecular nanomaterials.

Self-assembly of amphiphilic small molecules in water leads to nanostructures with customizable structure-property relationships arising from their tunable chemistries. Characterization of these assemblies is generally limited to their static structures -e.g. their geometries and dimensions - but the implementation of tools that provide a deeper understanding of molecular motions has recently emerged. Here, we summarize recent reports showcasing dynamics characterization tools and their application to small molecule assemblies, and we go on to highlight supramolecular systems whose properties are substantially affected by their conformational, exchange, and water dynamics. This review illustrates the importance of considering dynamics in rational amphiphile design.

A Global Minimization Toolkit for Batch-Fitting and χ2 Cluster Analysis of CW-EPR Spectra.

Electron paramagnetic resonance spectroscopy (EPR) is a uniquely powerful technique for characterizing conformational dynamics at specific sites within a broad range of molecular species in water. Computational tools for fitting EPR spectra have enabled dynamics parameters to be determined quantitatively. These tools have dramatically broadened the capabilities of EPR dynamics analysis, however, their implementation can easily lead to overfitting or problems with self-consistency. As a result, dynamics parameters and associated properties become difficult to reliably determine, particularly in the slow-motion regime. Here, we present an EPR analysis strategy and the corresponding computational tool for batch-fitting EPR spectra and cluster analysis of the χ2 landscape in Linux. We call this tool CSCA (Chi-Squared Cluster Analysis). The CSCA tool allows us to determine self-consistent rotational diffusion rates and enables calculations of activation energies of diffusion from Arrhenius plots. We demonstrate CSCA using a model system designed for EPR analysis: a self-assembled nanoribbon with radical electron spin labels positioned at known distances off the surface. We anticipate that the CSCA tool will increase the reproducibility of EPR fitting for the characterization of dynamics in biomolecules and soft matter.

Conformational Dynamics in Extended RGD-Containing Peptides.

RGD is a prolific example of a tripeptide used in biomaterials for cell adhesion, but the potency of free or surface-bound RGD tripeptide is orders-of-magnitude less than the RGD domain within natural proteins. We designed a set of peptides with varying lengths, composed of fragments of fibronectin protein whose central three residues are RGD, in order to vary their conformational behavior without changing the binding site's chemical environment. With these peptides, we measure the conformational dynamics and transient structure of the active site. Our studies reveal how flanking residues affect conformational behavior and integrin binding. We find that disorder of the binding site is important to the potency of RGD peptides and that transient hydrogen bonding near the RGD site affects both the energy landscape roughness of the peptides and peptide binding. This phenomenon is independent of longer-range folding interactions and helps explain why short binding sequences, including RGD itself, do not fully replicate the integrin-targeting properties of extracellular matrix proteins. Our studies reinforce that peptide binding is a holistic event and fragments larger than those directly involved in binding should be considered in the design of peptide epitopes for functional biomaterials.

Water Dynamics from the Surface to the Interior of a Supramolecular Nanostructure.

Water within and surrounding the structure of a biological system adopts context-specific dynamics that mediate virtually all of the events involved in the inner workings of a cell. These events range from protein folding and molecular recognition to the formation of hierarchical structures. Water dynamics are mediated by the chemistry and geometry of interfaces where water and biomolecules meet. Here we investigate experimentally and computationally the translational dynamics of vicinal water molecules within the volume of a supramolecular peptide nanofiber measuring 6.7 nm in diameter. Using Overhauser dynamic nuclear polarization relaxometry, we show that drastic differences exist in water motion within a distance of about one nanometer from the surface, with rapid diffusion in the hydrophobic interior and immobilized water on the nanofiber surface. These results demonstrate that water associated with materials designed at the nanoscale is not simply a solvent, but rather an integral part of their structure and potential functions.

Energy landscapes and functions of supramolecular systems.

By means of two supramolecular systems--peptide amphiphiles engaged in hydrogen-bonded ß-sheets, and chromophore amphiphiles driven to assemble by π-orbital overlaps--we show that the minima in the energy landscapes of supramolecular systems are defined by electrostatic repulsion and the ability of the dominant attractive forces to trap molecules in thermodynamically unfavourable configurations. These competing interactions can be selectively switched on and off, with the order of doing so determining the position of the final product in the energy landscape. Within the same energy landscape, the peptide-amphiphile system forms a thermodynamically favoured product characterized by long bundled fibres that promote biological cell adhesion and survival, and a metastable product characterized by short monodisperse fibres that interfere with adhesion and can lead to cell death. Our findings suggest that, in supramolecular systems, functions and energy landscapes are linked, superseding the more traditional connection between molecular design and function.

Internal dynamics of a supramolecular nanofibre.

A large variety of functional self-assembled supramolecular nanostructures have been reported over recent decades. The experimental approach to these systems initially focused on the design of molecules with specific interactions that lead to discrete geometric structures, and more recently on the kinetics and mechanistic pathways of self-assembly. However, there remains a major gap in our understanding of the internal conformational dynamics of these systems and of the links between their dynamics and function. Molecular dynamics simulations have yielded information on the molecular fluctuations of supramolecular assemblies, yet experimentally it has been difficult to obtain analogous data with subnanometre spatial resolution. Using site-directed spin labelling and electron paramagnetic resonance spectroscopy, we measured the conformational dynamics of a self-assembled nanofibre in water through its 6.7 nm cross-section. Our measurements provide unique insight for the design of supramolecular functional materials.

Asymmetric collapse in biomimetic complex coacervates revealed by local polymer and water dynamics.

Complex coacervation is a phenomenon characterized by the association of oppositely charged polyelectrolytes into micrometer-scale liquid condensates. This process is the purported first step in the formation of underwater adhesives by sessile marine organisms, as well as the process harnessed for the formation of new synthetic and protein-based contemporary materials. Efforts to understand the physical nature of complex coacervates are important for developing robust adhesives, injectable materials, or novel drug delivery vehicles for biomedical applications; however, their internal fluidity necessitates the use of in situ characterization strategies of their local dynamic properties, capabilities not offered by conventional techniques such as X-ray scattering, microscopy, or bulk rheological measurements. Herein, we employ the novel magnetic resonance technique Overhauser dynamic nuclear polarization enhanced nuclear magnetic resonance (DNP), together with electron paramagnetic resonance (EPR) line shape analysis, to concurrently quantify local molecular and hydration dynamics, with species- and site-specificity. We observe striking differences in the structure and dynamics of the protein-based biomimetic complex coacervates from their synthetic analogues, which is an asymmetric collapse of the polyelectrolyte constituents. From this study we suggest charge heterogeneity within a given polyelectrolyte chain to be an important parameter by which the internal structure of complex coacervates may be tuned. Acquiring molecular-level insight to the internal structure and dynamics of dynamic polymer complexes in water through the in situ characterization of site- and species-specific local polymer and hydration dynamics should be a promising general approach that has not been widely employed for materials characterization.

Self-Assembly for the Synthesis of Functional Biomaterials.

The use of self-assembly for the construction of functional biomaterials is a highly promising and exciting area of research, with great potential for the treatment of injury or disease. By using multiple noncovalent interactions, coded into the molecular design of the constituent components, self-assembly allows for the construction of complex, adaptable, and highly tunable materials with potent biological effects. This review describes some of the seminal advances in the use of self-assembly to make novel systems for regenerative medicine and biology. Materials based on peptides, proteins, DNA, or hybrids thereof have found application in the treatment of a wide range of injuries and diseases, and this review outlines the design principles and practical applications of these systems. Most of the examples covered focus on the synthesis of hydrogels for the scaffolding or transplantation of cells, with an emphasis on the biological, mechanical, and structural properties of the resulting materials. In addition, we will discuss the distinct advantages conferred by self-assembly (compared with traditional covalent materials), and present some of the challenges and opportunities for the next generation of self-assembled biomaterials.

Self-assembly of an optically active conjugated oligoelectrolyte.

Conjugated oligoelectrolytes are of emerging technological interest due to their recent function in the fabrication of optoelectronic devices, application in biosensors, and as species that facilitate transmembrane charge migration. Solubility in aqueous, or highly polar, solvents is important for many of these applications; however, there are few studies on how the self-assembly of conjugated oligoelectrolytes into multichromophore species influences linear and nonlinear optical properties. Here, we examine 1,4-bis(4'-(N,N-bis(6''-(N,N,N-trimethylammonium)hexyl)amino)-styryl)benzene tetraiodide (DSBNI) in water, a conjugated oligoelectrolyte based on the distyrylbenzene framework. We find that DSBNI aggregation leads to increased fluorescence lifetimes, coupled with hypsochromic shifts, and larger two-photon absorption cross sections. Liquid atomic force microscopy (AFM) and cryogenic transmission electron microscopy (cryo-TEM) were used to image DSBNI aggregates and to confirm that the planar molecules stack to form nanocylinders above a critical aggregation concentration. Finally, small-angle neutron scattering (SANS) was used to quantify the aggregate dimensions in situ. Comparison of the results highlights that the hydrophilic mica surface used to image via liquid AFM and the high concentrations required for cryo-TEM facilitate the propagation of the cylinders into long fibers. SANS experiments are consistent with equivalent molecular packing geometry but lower aspect ratios. It is therefore possible to understand the evolution of optical properties as a function of concentration and aggregation and the general geometric features of the resulting supramolecular structures.

Emerging investigator series: Aramid amphiphile nanoribbons for the remediation of lead from contaminated water.

Self-assembled nanoribbons from small molecule amphiphiles with chelating head groups and a structural domain to impart mechanical stability are reported for the remediation of lead from contaminated water. The nanoribbons' remediation capacity is affected by pH and the presence of competing cations, and can be modulated by head group choice.

Domain-selective thermal decomposition within supramolecular nanoribbons.

Self-assembly of small molecules in water provides a powerful route to nanostructures with pristine molecular organization and small dimensions (<10 nm). Such assemblies represent emerging high surface area nanomaterials, customizable for biomedical and energy applications. However, to exploit self-assembly, the constituent molecules must be sufficiently amphiphilic and satisfy prescribed packing criteria, dramatically limiting the range of surface chemistries achievable. Here, we design supramolecular nanoribbons that contain: (1) inert and stable internal domains, and (2) sacrificial surface groups that are thermally labile, and we demonstrate complete thermal decomposition of the nanoribbon surfaces. After heating, the remainder of each constituent molecule is kinetically trapped, nanoribbon morphology and internal organization are maintained, and the nanoribbons are fully hydrophobic. This approach represents a pathway to form nanostructures that circumvent amphiphilicity and packing parameter constraints and generates structures that are not achievable by self-assembly alone, nor top-down approaches, broadening the utility of molecular nanomaterials for new targets.

Self-assembly of aramid amphiphiles into ultra-stable nanoribbons and aligned nanoribbon threads.

Small-molecule self-assembly is an established route for producing high-surface-area nanostructures with readily customizable chemistries and precise molecular organization. However, these structures are fragile, exhibiting molecular exchange, migration and rearrangement-among other dynamic instabilities-and are prone to dissociation upon drying. Here we show a small-molecule platform, the aramid amphiphile, that overcomes these dynamic instabilities by incorporating a Kevlar-inspired domain into the molecular structure. Strong, anisotropic interactions between aramid amphiphiles suppress molecular exchange and elicit spontaneous self-assembly in water to form nanoribbons with lengths of up to 20 micrometres. Individual nanoribbons have a Young's modulus of 1.7 GPa and tensile strength of 1.9 GPa. We exploit this stability to extend small-molecule self-assembly to hierarchically ordered macroscopic materials outside of solvated environments. Through an aqueous shear alignment process, we organize aramid amphiphile nanoribbons into arbitrarily long, flexible threads that support 200 times their weight when dried. Tensile tests of the dry threads provide a benchmark for Young's moduli (between ~400 and 600 MPa) and extensibilities (between ~0.6 and 1.1%) that depend on the counterion chemistry. This bottom-up approach to macroscopic materials could benefit solid-state applications historically inaccessible by self-assembled nanomaterials.

Quantifying residue-specific conformational dynamics of a highly reactive 29-mer peptide.

Understanding structural transitions within macromolecules remains an important challenge in biochemistry, with important implications for drug development and medicine. Insight into molecular behavior often requires residue-specific dynamics measurement at micromolar concentrations. We studied MP01-Gen4, a library peptide selected to rapidly undergo bioconjugation, by using electron paramagnetic resonance (EPR) to measure conformational dynamics. We mapped the dynamics of MP01-Gen4 with residue-specificity and identified the regions involved in a structural transformation related to the conjugation reaction. Upon reaction, the conformational dynamics of residues near the termini slow significantly more than central residues, indicating that the reaction induces a structural transition far from the reaction site. Arrhenius analysis demonstrates a nearly threefold decrease in the activation energy of conformational diffusion upon reaction (8.0 kBT to 3.4 kBT), which occurs across the entire peptide, independently of residue position. This novel approach to EPR spectral analysis provides insight into the positional extent of disorder and the nature of the energy landscape of a highly reactive, intrinsically disordered library peptide before and after conjugation.

Beyond Covalent Crosslinks: Applications of Supramolecular Gels.

Traditionally, gels have been defined by their covalently cross-linked polymer networks. Supramolecular gels challenge this framework by relying on non-covalent interactions for self-organization into hierarchical structures. This class of materials offers a variety of novel and exciting potential applications. This review draws together recent advances in supramolecular gels with an emphasis on their proposed uses as optoelectronic, energy, biomedical, and biological materials. Additional special topics reviewed include environmental remediation, participation in synthesis procedures, and other industrial uses. The examples presented here demonstrate unique benefits of supramolecular gels, including tunability, processability, and self-healing capability, enabling a new approach to solve engineering challenges.

Cell death versus cell survival instructed by supramolecular cohesion of nanostructures.

Many naturally occurring peptides containing cationic and hydrophobic domains have evolved to interact with mammalian cell membranes and have been incorporated into materials for non-viral gene delivery, cancer therapy or treatment of microbial infections. Their electrostatic attraction to the negatively charged cell surface and hydrophobic interactions with the membrane lipids enable intracellular delivery or cell lysis. Although the effects of hydrophobicity and cationic charge of soluble molecules on the cell membrane are well known, the interactions between materials with these molecular features and cells remain poorly understood. Here we report that varying the cohesive forces within nanofibres of supramolecular materials with nearly identical cationic and hydrophobic structure instruct cell death or cell survival. Weak intermolecular bonds promote cell death through disruption of lipid membranes, while materials reinforced by hydrogen bonds support cell viability. These findings provide new strategies to design biomaterials that interact with the cell membrane.