Domain Swapping in Abiotic Foldamers.

Foldamer sequences that adopt tertiary helix-turn-helix folds mediated by helix-helix hydrogen bonding in organic solvents have been previously reported. In an attempt to create genuine abiotic quaternary structures, i.e. assemblies of tertiary structures, new sequences were prepared that possess additional hydrogen bond donors at positions that may promote an association between the tertiary folds. However, a solid state structure and extensive solution state investigations by Nuclear Magnetic Resonance (NMR) and Circular Dichroism (CD) show that, instead of forming a quaternary structure, the tertiary folds assemble into stable domain-swapped dimer motifs. Domain swapping entails a complete reorganization of the arrays of hydrogen bonds and changes in relative helix orientation and handedness that can all be rationalized.

Molecular Sensing and Manipulation of Protein Oligomerization in Membrane Nanotubes with Bolaamphiphilic Foldamers.

Adaptive and reversible self-assembly of supramolecular protein structures is a fundamental characteristic of dynamic living matter. However, the quantitative detection and assessment of the emergence of mesoscale protein complexes from small and dynamic oligomeric precursors remains highly challenging. Here, we present a novel approach utilizing a short membrane nanotube (sNT) pulled from a planar membrane reservoir as nanotemplates for molecular reconstruction, manipulation, and sensing of protein oligomerization and self-assembly at the mesoscale. The sNT reports changes in membrane shape and rigidity caused by membrane-bound proteins as variations of the ionic conductivity of the sNT lumen. To confine oligomerization to the sNT, we have designed and synthesized rigid oligoamide foldamer tapes (ROFTs). Charged ROFTs incorporate into the planar and sNT membranes, mediate protein binding to the membranes, and, driven by the luminal electric field, shuttle the bound proteins between the sNT and planar membranes. Using Annexin-V (AnV) as a prototype, we show that the sNT detects AnV oligomers shuttled into the nanotube by ROFTs. Accumulation of AnV on the sNT induces its self-assembly into a curved lattice, restricting the sNT geometry and inhibiting the material uptake from the reservoir during the sNT extension, leading to the sNT fission. By comparing the spontaneous and ROFT-mediated entry of AnV into the sNT, we reveal how intricate membrane curvature sensing by small AnV oligomers controls the lattice self-assembly. These results establish sNT-ROFT as a powerful tool for molecular reconstruction and functional analyses of protein oligomerization and self-assembly, with broad application to various membrane processes.

An abiotic, tetrameric, eight-helix bundle.

Four helically folded aromatic oligoamide sequences containing either a chiral monomer based on 2-(2-aminophenoxy)-propionic acid, an N-terminal (1H)-camphanyl group, or both, were synthesized. Spectroscopic solution investigations using 1H NMR and circular dichroism (CD) demonstrated that the 2-(2-aminophenoxy)-propionic acid unit biases helix handedness quantitatively in chloroform and dichloromethane. It even quantitatively overcomes an opposing effect of the camphanyl group and thus ensures reliable helix handedness control. A series of nine sequences composed of two helically folded aromatic oligoamide segments separated by a flexible linker based on a di-, tri- or tetraethylene glycol unit were then synthesized. In these sequences, helix handedness was controlled by means of an N-terminal (1H)-camphanyl group or a 2-(2-aminophenoxy)-propionic acid units in either both helical segments, or only in the N-terminal segment, or in none of the segments. The helical segments all displayed hydroxy and carbonyl groups at their surfaces as hydrogen bond donors and acceptors so as to promote helix-to-helix hydrogen bonding. NMR and CD spectroscopic studies showed that, in some cases, well-defined, stable, discrete abiotic helix-turn-helix tertiary folds form in organic solvents. Molecular modelling suggests that these correspond to structures in which the two helix axes are at an angle. In one case, the absence of handedness control resulted in a complex and large aggregate. A solid state structure obtained by single crystal X-ray diffraction analysis revealed a tetrameric assembly composed of eight helices with both right and left handedness arranged in three subdomains consisting of two hydrogen-bonded three-helix bundles and one two-helix-bundle. Several helix-to-helix hydrogen bonds were mediated by bridging water molecules. This structure constitutes an important milestone in the construction of abiotic protein-like architectures.

Coaxial assembly of helical aromatic foldamers by metal coordination.

Deprotonation of acid-terminated helical aromatic foldamers with a mineral base in chlorinated solvents led to their dimerization through the coordination of a metal cation (Li+, Na+, K+, Ag+, or Hg2+) with the terminal carboxylate functions. This new ligation method was applied to oligomerize diacid-functionalized foldamers.

Homochiral versus Heterochiral Dimeric Helical Foldamer Bundles: Chlorinated-Solvent-Dependent Self-Sorting.

Aromatic oligoamide sequences programmed to fold into stable helical conformations were designed to display a linear array of hydrogen-bond donors and acceptors at their surface. Sequences were prepared by solid-phase synthesis. Solution 1 H NMR spectroscopic studies and solid-state crystallographic structures demonstrated the formation of stable hydrogen-bond-mediated dimeric helix bundles that could be either heterochiral (with a P and an M helix) or homochiral (with two P or two M helices). Formation of the hetero- or homochiral dimers could be driven quantitatively using different chlorinated solvents-exemplifying a remarkable case of either social or narcissistic chiral self-sorting or upon imposing absolute handedness to the helices to forbid PM species.

Molecular torsion springs: alteration of helix curvature in frustrated tertiary folds.

The first abiotic foldamer tertiary structures have been recently reported in the form of aromatic helix-turn-helix motifs based on oligo-quinolinecarboxamides held together by intramolecular hydrogen bonds. Tertiary folds were predicted by computational modelling of the hydrogen-bonding interfaces between helices and later verified by X-ray crystallography. However, the prognosis of how the conformational preference inherent to each helix influences the tertiary structure warranted further investigation. Several new helix-turn-helix sequences were synthesised in which some hydrogen bonds have been removed. Contrary to expectations, this change did not strongly destabilise the tertiary folds. On closer inspection, a new crystal structure revealed that helices adopt their natural curvature when some hydrogen bonds are missing and undergo some spring torsion upon forming the said hydrogen bonds, thus potentially giving rise to a conformational frustration. This phenomenon sheds light on the aggregation behaviour of the helices when they are not linked by a turn unit.

Directing the Self-Assembly of Aromatic Foldamer Helices using Acridine Appendages and Metal Coordination.

Folded molecules provide complex interaction interfaces amenable to sophisticated self-assembly motifs. Because of their high conformational stability, aromatic foldamers constitute suitable candidates for the rational elaboration of self-assembled architectures. Several multiturn helical aromatic oligoamides have been synthesized that possess arrays of acridine appendages pointing in one or two directions. The acridine units were shown to direct self-assembly in the solid state via aromatic stacking leading to recurrent helix-helix association patterns under the form of discrete dimers or extended arrays. In the presence of Pd(II), metal coordination of the acridine units overwhelms other forces and generates new metal-mediated multihelical self-assemblies, including macrocycles. These observations demonstrate simple access to different types of foldamer-containing architectures, ranging from discrete objects to 1D and, by extension, 2D and 3D arrays.

Structural adaptations of electrosprayed aromatic oligoamide foldamers on Ag(111).

Aromatic foldamers are promising for applications such as molecular recognition and molecular machinery. For many of these, defect free, 2D-crystaline monolayers are needed. To this end, submonolayers were prepared in ultra-high vacuum (UHV) on Ag(111) via electrospray controlled ion beam deposition (ES-CIBD). On the surface, the unfolded state is unambiguously identified by real-space single-molecule imaging using scanning tunnelling microscopy (STM) and it is found to assemble in regular structures.

Selective and Cooperative Photocycloadditions within Multistranded Aromatic Sheets.

A series of aromatic helix-sheet-helix oligoamide foldamers composed of several different photosensitive diazaanthracene units have been designed and synthesized. Molecular objects up to 7 kDa were straightforwardly produced on a 100 mg scale. Nuclear magnetic resonance and crystallographic investigations revealed that helix-sheet-helix architectures can adopt one or two distinct conformations. Sequences composed of an even number of turn units were found to fold in a canonical symmetrical conformation with two helices of identical handedness stacked above and below the sheet segment. Sequences composed of an odd number of turns revealed a coexistence between a canonical fold with helices of opposite handedness and an alternate fold with a twist within the sheet and two helices of identical handedness. The proportions between these species could be manipulated, in some cases quantitatively, being dependent on solvent, temperature, and absolute control of helix handedness. Diazaanthracene units were shown to display distinct reactivity toward [4 + 4] photocycloadditions according to the substituent in position 9. Their organization within the sequences was programmed to allow photoreactions to take place in a specific order. Reaction pathways and kinetics were deciphered and product characterized, demonstrating the possibility to orchestrate successive photoreactions so as to avoid orphan units or to deliberately produce orphan units at precise locations. Strong cooperative effects were observed in which the photoreaction rate was influenced by the presence (or absence) of photoadducts in the structure. Multiple photoreactions within the aromatic sheet eventually lead to structure lengthening and stiffening, locking conformational equilibria. Photoproducts could be thermally reverted.

Loading Linear Arrays of CuII Inside Aromatic Amide Helices.

The very stable helices of 8-amino-2-quinolinecarboxylic acid oligoamides are shown to uptake CuII ions in their cavity through deprotonation of their amide functions with minimal alteration of their shape, unlike most metallo-organic structures which generally differ from their organic precursors. The outcome is the formation of intramolecular linear arrays of a defined number of CuII centers (up to sixteen in this study) at a 3 Šdistance, forming a molecular mimic of a metal wire completely surrounded by an organic sheath. The helices pack in the solid state so that the arrays of CuII extend intermolecularly. Conductive-AFM and cyclic voltammetry suggest that electrons are transported throughout the metal-loaded helices in contrast with hole transport observed for analogous foldamers devoid of metal ions.

Oligo-Quinolylene-Vinylene Foldamers.

Quinoline based aromatic amide foldamers are known to adopt stable folded conformations. We have developed a synthetic approach to produce similar oligomers where all amide bonds, or part of them, have been replaced by an isosteric vinylene group. The results of solution and solid state structural studies show that oligomers exclusively containing vinylene linkages are not well folded, and adopt predominantly flat conformations. In contrast, a vinylene segment flanked by helical oligoamides also folds in a helix, albeit with a slightly lower curvature. The presence of vinylene functions also result in an extension of π-conjugation across the oligomer that may change charge transport properties. Altogether, these results pave the way to foldamers in which both structural control and specific electronic properties may be engineered.

Large-Amplitude Conformational Changes in Self-Assembled Multi-Stranded Aromatic Sheets.

The orchestration of ever larger conformational changes is made possible by the development of increasingly complex foldamers. Aromatic sheets, a rare motif in synthetic foldamer structures, have been designed so as to form discrete stacks of intercalated aromatic strands through the self-assembly of two identical subunits. Ion-mobility ESI-MS confirms the formation of compact dimers. X-ray crystallography reveals the existence of two distinct conformational dimeric states that require large changes to interconvert. Molecular dynamics simulation validates the stability of the two conformations and the possibility of their interconversion.

Mussel-Inspired Dual-Cross-linking Hyaluronic Acid/ε-Polylysine Hydrogel with Self-Healing and Antibacterial Properties for Wound Healing.

Physicians have long been calling for an inherent antimicrobial wound dressing, which will be a great progress for treating complicated infections. Here, we report a novel bioadhesive hydrogel with inherent antibacterial properties prepared by mixing modified hyaluronic acid (HA) and ε-polylysine (EPL). This hydrogel can effectively kill Gram (+) and (-) bacteria for its high positive charge density on the surface. The sol-gel transition occurs within seconds via horseradish peroxidase enzymatic cross-linking and Schiff base reaction, which also allows the hydrogel to recover completely from destruction quickly within 5 min. In an infected rat wound model, histological studies indicated that the hydrogels effectively killed bacteria on the surface of wounds and accelerated wound healing. Histological analysis indicated that the thickness of the newborn skin, the density of the newborn microvascular, granulation tissue, and the collagen of rats treated with hydrogel dressings were twice as high as those treated by commercial fibrin glue. These results indicate that the HA/EPL hydrogel has great potential as an antibacterial wound dressing for future clinical applications.

Parallel Homochiral and Anti-Parallel Heterochiral Hydrogen-Bonding Interfaces in Multi-Helical Abiotic Foldamers.

A hydrogen-bonding interface between helical aromatic oligoamide foldamers has been designed to promote the folding of a helix-turn-helix motif with a head-to-tail arrangement of two helices of opposite handedness. This design complements an earlier helix-turn-helix motif with a head-to-head arrangement of two helices of identical handedness interface. The two motifs were shown to have comparable stability and were combined in a unimolecular tetra-helix fold constituting the largest abiotic tertiary structure to date.

Simplification in the Acquisition and Analysis of Fluorescence Decays Acquired with Polarized Emission for Time-Resolved Fluorescence Anisotropy Measurements.

This study introduces a global fluorescence decay analysis that substantially simplifies the acquisition and analysis of time-resolved fluorescence decays acquired with a vertically polarized excitation and vertically (IVV(t)) and horizontally (IVH(t)) polarized emission for time-resolved fluorescence anisotropy (TRFA) measurements. TRFA measurements were conducted whereby the IVV(t) and IVH(t) fluorescence decays of a series of oligoquinolines labeled at one end with an oligo(phenylenevinylene) dye (OPV-Qn with n = 4, 7, 17, 24, 33) were acquired according to the standard protocol that is currently accepted in the scientific literature which involves toggling the emission polarizer before fitting linear combinations of the IVV(t) and IVH(t) decays or acquiring the IVV(t) and IVH(t) decays with static polarizers before fitting them globally. The rotational time (ϕ) and initial anisotropy (r0) retrieved from these analyses were identical within experimental error regardless of whether the decays were acquired with toggling or static polarizers and fitted according to the standard protocol or globally. These experimental results were further supported by retrieving the parameters used to generate mono-, bi-, and tri-exponential TRFAs from the global analysis of simulated IVV(t) and IVH(t) fluorescence decays which were found to match perfectly the values that were inputted. Together, these experiments and simulations demonstrated that the parameters describing any type of TRFA can be extracted directly from the analysis of the IVV(t) and IVH(t) fluorescence decays acquired with a standard time-resolved fluorometer, a substantial simplification compared to the protocols currently in place to determine the TRFA.

Interplay of secondary and tertiary folding in abiotic foldamers.

The first true abiotic tertiary folded structures, i.e. at the exclusion of any aliphatic amino acid, have recently been introduced under the form of aromatic oligoamide helix-turn-helix foldamers stabilized by hydrogen bonds in organic solvents. We present an investigation of the interplay of secondary and tertiary folding and of some cooperative effects in these systems. A solid phase synthesis approach to the preparation of these sequences was developed to facilitate systematic variation. Flexible pyridine-based units were introduced in various proportions in replacement of more rigid quinoline-based units. Conformational behaviour was assessed in solution by NMR, in the solid state by X-ray crystallography, and computationally through molecular dynamics simulations. Altogether, our results demonstrate that tertiary folding stabilizes otherwise flexible secondary structures, and that the disruption of tertiary folds upon adding polar solvents follows different mechanisms depending on whether secondary structures are inherently stable or not. These findings constitute a solid basis on which to further increase the size and complexity of abiotic folded structures and to eventually orchestrate folding dynamics and responsiveness.

Aromatic ß-sheet foldamers based on tertiary squaramides.

The preference of N,N-aryl, alkyl tertiary amides for cis conformations has been exploited through the use of tertiary squaramides as hairpin turn units that promote the folding of aromatic ß-sheets. Head-to-head aromatic arrangements were shown to prevail in sufficiently long bent aromatic sequences.

Light-Controlled Conformational Switch of an Aromatic Oligoamide Foldamer.

An aromatic oligoamide sequence composed of a light-responsive diazaanthracene-based aromatic ß-sheet flanked by two variable diameter helical segments was prepared. Structural investigations revealed that such oligomers adopt two distinct conformations: a canonical symmetrical conformation with the two helices stacked above and below the sheet, and an unanticipated unsymmetrical conformation in which one helix has flipped to directly stack with the first helix. Photoirradiation of the foldamer led to the quantitative, and thermally reversible, formation of a single photoproduct resulting from the [4+4] cycloaddition of two diazaanthracenes within the aromatic ß-sheet. NMR and crystallographic studies revealed a parallel arrangement of the diazaanthracene photoproduct and a complete conversion into a symmetrical conformation requiring a rearrangement of all unsymmetrical conformers. These results highlight the potential of foldamers, with structures more complex than isolated helices, for the design of photoswitches showing nontrivial nanometer scale shape changes.

Designing cooperatively folded abiotic uni- and multimolecular helix bundles.

Abiotic foldamers, that is foldamers that have backbones chemically remote from peptidic and nucleotidic skeletons, may give access to shapes and functions different to those of peptides and nucleotides. However, design methodologies towards abiotic tertiary and quaternary structures are yet to be developed. Here we report rationally designed interactional patterns to guide the folding and assembly of abiotic helix bundles. Computational design facilitated the introduction of hydrogen-bonding functionalities at defined locations on the aromatic amide backbones that promote cooperative folding into helix-turn-helix motifs in organic solvents. The hydrogen-bond-directed aggregation of helices not linked by a turn unit produced several thermodynamically and kinetically stable homochiral dimeric and trimeric bundles with structures that are distinct from the designed helix-turn-helix. Relative helix orientation within the bundles may be changed from parallel to tilted on subtle solvent variations. Altogether, these results prefigure the richness and uniqueness of abiotic tertiary structure behaviour.

Multi-dimensional charge transport in supramolecular helical foldamer assemblies.

Aromatic foldamers are bioinspired architectures whose potential use in materials remains largely unexplored. Here we report our investigation of vertical and horizontal charge transport over long distances in helical oligo-quinolinecarboxamide foldamers organized as single monolayers on Au or SiO2. Conductive atomic force microscopy showed that vertical conductivity is efficient and that it displays a low attenuation with foldamer length (0.06 Å-1). In contrast, horizontal charge transport is found to be negligible, demonstrating the strong anisotropy of foldamer monolayers. Kinetic Monte Carlo calculations were used to probe the mechanism of charge transport in these helical molecules and revealed the presence of intramolecular through-space charge transfer integrals approaching those found in pentacene and rubrene crystals, in line with experimental results. Kinetic Monte Carlo simulations of charge hopping along the foldamer chain evidence the strong contribution of multiple 1D and 3D pathways in these architectures and their dependence on conformational order. These findings show that helical foldamer architectures may provide a route for achieving charge transport over long distance by combining multiple charge transport pathways.