A Combination of Bio-Orthogonal Supramolecular Clicking and Proximity Chemical Tagging as a Supramolecular Tool for Discovery of Putative Proteins Associated with Laminopathic Disease.

Protein mutations alter protein-protein interactions that can lead to a number of illnesses. Mutations in lamin A (LMNA) have been reported to cause laminopathies. However, the proteins associated with the LMNA mutation have mostly remained unexplored. Herein, a new chemical tool for proximal proteomics is reported, developed by a combination of proximity chemical tagging and a bio-orthogonal supramolecular latching based on cucurbit[7]uril (CB[7])-based host-guest interactions. As this host-guest interaction acts as a noncovalent clickable motif that can be unclicked on-demand, this new chemical tool is exploited for reliable detection of the proximal proteins of LMNA and its mutant that causes laminopathic dilated cardiomyopathy (DCM). Most importantly, a comparison study reveals, for the first time, mutant-dependent alteration in LMNA proteomic environments, which allows to identify putative laminopathic DCM-linked proteins including FOXJ3 and CELF2. This study demonstrates the feasibility of this chemical tool for reliable proximal proteomics, and its immense potential as a new research platform for discovering biomarkers associated with protein mutation-linked diseases.

Flow Synthesis of Gigantic Porphyrinic Cages: Facile Synthesis of P12 L24 and Discovery of Kinetic Product P9 L18.

A continuous flow methodology for the facile and high-yielding synthesis of the porphyrin-based self-assembled organic cage, P12 L24 is reported, along with the serendipitous discovery of a kinetic product, P9 L18 cage, which has been characterized by MALDI-TOF MS, NMR, and AFM analysis. A theoretical study suggests a tricapped trigonal prismatic geometry for P9 L18 . Unlike P12 L24 , P9 L18 is unstable and readily decomposes into monomers and small oligomers. While the batch synthesis produces only the thermodynamic product P12 L24 , the continuous flow process generates not only the thermodynamic product but also kinetic products, such as P9 L18 , illustrating the advantages of the continuous flow process for the synthesis of self-assembled cages and the exploration of new non-equilibrium assemblies.

Protomer of Imipramine Captured in Cucurbit[7]uril.

Small molecules possessing multiple proton-accessible sites are important not only to many biological systems but also to host-guest chemistry; their protonation states are causal to boosting or hindering specific host-guest interactions. However, determining the protonation site is not trivial. Herein, we conduct electrospray ionization ion mobility spectrometry-mass spectrometry to imipramine, a known molecule with two protonation sites, based on the introduction of cucurbit[7]uril as a host molecule. For protonated imipramine, the proposed strategy allows clear distinction of the two protomers as host-guest complex ions and can be leveraged to capture the energetically less preferable protomer of the protonated imipramine.

Synthetic Monosaccharide Channels: Size-Selective Transmembrane Transport of Glucose and Fructose Mediated by Porphyrin Boxes.

Here we report synthetic monosaccharide channels built with shape-persistent organic cages, porphyrin boxes (PBs), that allow facile transmembrane transport of glucose and fructose through their windows. PBs show a much higher transport rate for glucose and fructose over disaccharides such as sucrose, as evidenced by intravesicular enzyme assays and molecular dynamics simulations. The transport rate can be modulated by changing the length of the alkyl chains decorating the cage windows. Insertion of a linear pillar ligand into the cavity of PBs blocks the monosaccharide transport. In vitro cell experiment shows that PBs transport glucose across the living-cell membrane and enhance cell viability when the natural glucose transporter GLUT1 is blocked. Time-dependent live-cell imaging and MTT assays confirm the cyto-compatibility of PBs. The monosaccharide-selective transport ability of PBs is reminiscent of natural glucose transporters (GLUTs), which are crucial for numerous biological functions.

Contagious Aggregation: Transmittable Protein Aggregation in Cellular Communities Initiated by Synthetic Cells.

Aggregation of amyloidogenic proteins causing neurodegenerative diseases is an uncontrollable and contagious process that is often associated with lipid membranes in a highly complex physiological environment. Although several approaches using natural cells and membrane models have been reported, systematic investigations focusing on the association with the membranes are highly challenging, mostly because of the lack of proper molecular tools. Here, we report a new supramolecular approach using a synthetic cell system capable of controlling the initiation of protein aggregation and mimicking various conditions of lipid membranes, thereby enabling systematic investigations of membrane-dependent effects on protein aggregation by visualization. Extending this strategy through concurrent use of synthetic cells and natural cells, we demonstrate the potential of this approach for systematic and in-depth studies on interrogating inter- and intracellularly transmittable protein aggregation. Thus, this new approach offers opportunities for gaining insights into the pathological implications of contagious protein aggregation associated with membranes for neurotoxicity.

Dramatically Enhanced Reactivity of Fullerenes and Tetrazine towards the Inverse-Electron-Demand Diels-Alder Reaction inside a Porous Porphyrinic Cage.

Inverse-electron-demand Diels-Alder reaction (IEDDA) between fullerenes and 1,2,4,5-tetrazine generally requires harsh conditions and long reaction times due to their strong electron-accepting nature. Herein, we report a dramatic enhancement in the reactivity of the fullerenes (C60 /C70 )-tetrazine reaction inside a porous Zn-porphyrinic cage (Zn-PB) under sustainable conditions by installing a tetrazine-based axle (LA) via metal-ligand coordination bond, which modulates the cavity size to facilitate the encapsulation of fullerenes. Upon encapsulation, the close proximity of fullerenes and the tetrazine group of LA dramatically increase their reactivity towards the IEDDA reaction to form fullerene-tetrazine adducts. Furthermore, the C60 -tetrazine adduct is rearranged upon hydration to a bent-shaped C60 -pyrazoline adduct that can be released from the Zn-PB cavity in the presence of excess LA, thus catalyzing the formation of C60 -pyrazoline adduct inside Zn-PB without product inhibition.

Lipid-Oriented Live-Cell Distinction of B and T Lymphocytes.

The identification of each cell type is essential for understanding multicellular communities. Antibodies set as biomarkers have been the main toolbox for cell-type recognition, and chemical probes are emerging surrogates. Herein we report the first small-molecule probe, CDgB, to discriminate B lymphocytes from T lymphocytes, which was previously impossible without the help of antibodies. Through the study of the origin of cell specificity, we discovered an unexpected novel mechanism of membrane-oriented live-cell distinction. B cells maintain higher flexibility in their cell membrane than T cells and accumulate the lipid-like probe CDgB more preferably. Because B and T cells share common ancestors, we tracked the cell membrane changes of the progenitor cells and disclosed the dynamic reorganization of the membrane properties over the lymphocyte differentiation progress. This study casts an orthogonal strategy for the small-molecule cell identifier and enriches the toolbox for live-cell distinction from complex cell communities.

Construction of Stable Metal-Organic Framework Platforms Embedding N-Heterocyclic Carbene Metal Complexes for Selective Catalysis.

We report a bottom-up approach to immobilize catalysts into MOFs, including copper halides and gold chloride in a predictable manner. Interestingly, the structures of MOFs bearing NHC metal complexes maintained a similar 4-fold interpenetrated cube. They exhibited exceptionally high porosity despite the interpenetrated structure and showed good stability in various solvents. Moreover, these MOFs possess high size activity depending on the size of the substrates in various reactions, compared to homogeneous catalysis. Also, the high catalytic activity of MOFs can be preserved 4 times without significant loss of crystallinity. Incorporation of the various metal complexes into MOFs allows for the preparation of functional MOFs for practical applications.

Programmable Synthesis of Silver Wheels.

The synthesis of sandwich-shaped multinuclear silver complexes with planar penta- and tetranuclear wheel-shaped silver units and a central anion, [Agn(2-HPB)2(A-)](OTf-)n-1, nAgA, n = 4 or 5 and A- = OH- or F- or Cl-, is reported, along with complete spectroscopic and structural characterization. An NMR mechanistic study reveals that silver complexes were formed in the following order: 2Ag → 3AgH2O → 5AgOH → 4AgOH. The central hydroxides in 4AgOH and 5AgOH exhibit exotic physical properties due to the confined environment inside the complex. The size of these silver wheels can be tuned by changing the central anion or extracting/adding one silver atom. This study provides the facile way to synthesize discrete wheel-shaped multinuclear silver complexes and provides valuable insights into the dynamics of the self-assembly process.

Supramolecular Fullerene Tetramers Concocted with Porphyrin Boxes Enable Efficient Charge Separation and Delocalization.

Herein, we report a novel porphyrin/fullerene supramolecular cocrystal using a shape-persistent zinc-metalated porphyrin box (Zn-PB) and C60/C70. An unprecedented arrangement of a tightly packed square-planar core of four C60 or C70 surrounded by six cube-shaped Zn-PBs was observed. This unique packing promotes strong charge transfer (CT) interactions between the two components in the ground state and formation of charge-separated states with very long lifetimes in the excited state and enables unusually high photoconductivity. Quantum chemical calculations show that these features are enabled by delocalized orbitals that promote the CT, on one hand, and that are spatially separated from each other, on the other hand. This work may open a new avenue to design novel electron donor/acceptor architectures for artificial photosynthesis.

Supramolecular Tuning Enables Selective Oxygen Reduction Catalyzed by Cobalt Porphyrins for Direct Electrosynthesis of Hydrogen Peroxide.

We report a supramolecular strategy for promoting the selective reduction of O2 for direct electrosynthesis of H2 O2 . We utilized cobalt tetraphenylporphyrin (Co-TPP), an oxygen reduction reaction (ORR) catalyst with highly variable product selectivity, as a building block to assemble the permanently porous supramolecular cage Co-PB-1(6) bearing six Co-TPP subunits connected through twenty-four imine bonds. Reduction of these imine linkers to amines yields the more flexible cage Co-rPB-1(6). Both Co-PB-1(6) and Co-rPB-1(6) cages produce 90-100 % H2 O2 from electrochemical ORR catalysis in neutral pH water, whereas the Co-TPP monomer gives a 50 % mixture of H2 O2 and H2 O. Bimolecular pathways have been implicated in facilitating H2 O formation, therefore, we attribute this high H2 O2 selectivity to site isolation of the discrete molecular units in each supramolecule. The ability to control reaction selectivity in supramolecular structures beyond traditional host-guest interactions offers new opportunities for designing such architectures for a broader range of catalytic applications.

Hierarchical Self-Assembly of Poly-Pseudorotaxanes into Artificial Microtubules.

Hierarchical self-assembly of building blocks over multiple length scales is ubiquitous in living organisms. Microtubules are one of the principal cellular components formed by hierarchical self-assembly of nanometer-sized tubulin heterodimers into protofilaments, which then associate to form micron-length-scale, multi-stranded tubes. This peculiar biological process is now mimicked with a fully synthetic molecule, which forms a 1:1 host-guest complex with cucurbit[7]uril as a globular building block, and then polymerizes into linear poly-pseudorotaxanes that associate laterally with each other in a self-shape-complementary manner to form a tubular structure with a length over tens of micrometers. Molecular dynamic simulations suggest that the tubular assembly consists of eight poly-pseudorotaxanes that wind together to form a 4.5 nm wide multi-stranded tubule.

Superacid-Mediated Functionalization of Hydroxylated Cucurbit[n]urils.

Herein we report a facile transformation of hydroxylated cucurbit[n]uril (CB[n], n = 6 and 7) to other functionality-conjugated CB[n]s by nucleophilic substitution of the hydroxyl group with a wide range of nitriles and alcohols. The reaction proceeds efficiently via generation of a superelectrophilic carbocation on the CB framework from hydroxylated CB[n]s under superacidic conditions. One of the resulting CB[n] derivatives with reactive functionality, monocarboxylated CB[7], is efficiently conjugated to an enzyme (horseradish peroxidase, HRP) by amide coupling. This provides a CB[7]-conjugated functional biomaterial (CB[7]-HRP) that selectively detects proteins labeled with a guest, adamantylammonium (AdA), based on bioorthogonal high-affinity host-guest interactions between CB[7] and AdA. We demonstrated the potential of overcoming the limitations in preparing reactive functional CB[n] derivatives, enabling the exploration of novel bioapplications of CB[n]-based host-guest chemistry with new CB[n]-conjugated functional materials.

Porphyrin Boxes.

In order to fabricate efficient molecular photonic devices, it has been a long-held aspiration for chemists to understand and mimic natural light-harvesting complexes where a rapid and efficient transfer of excitation energy between chlorophyll pigments is observed. Synthetic porphyrins are attractive building blocks in this regard because of their rigid and planar geometry, high thermal and electronic stability, high molar extinction, small and tunable band gap, and tweakable optical as well as redox behavior. Owing to these fascinating properties, various types of porphyrin-based architectures have been reported utilizing both covalent and noncovalent approaches. However, it still remains a challenge to construct chemically robust, well-defined three-dimensional porphyrin cages which can be easily synthesized and yet suitable for useful applications both in solution as well as in solid state. Working on this idea, we recently synthesized box-shaped organic cages, which we called porphyrin boxes, by making use of dynamic covalent chemistry of imine condensation reaction between 4-connecting, square-shaped, tetraformylporphyrin and 3-connecting, triangular-shaped, triamine molecules. Various presynthetic, as well as postsynthetic modifications, can be carried out on porphyrin boxes including a variation of the alkyl chain length in their 3-connecting subunit, chemical functionalization, and metalation of the porphyrin core. This can remarkably tune their inherent properties, e.g., solubility, window size, volume, and polarity of the internal void. The porphyrin boxes can therefore be considered as a significant addition to the family of multiporphyrin-based architectures, and because of their chemical stability and shape persistency, the applications of porphyrin boxes expand beyond the photophysical properties of an artificial light-harvesting complex. Consequently, they have been exploited as porous organic cages, where their gas adsorption properties have been investigated. By incorporating them in a lipid bilayer membrane, an iodide selective synthetic ion channel has also been demonstrated. Further, we have explored electrocatalytic reduction of carbon dioxide using Fe(III) metalated porphyrin boxes. Additionally, the precise size and ease of metalation of porphyrin boxes allowed us to utilize them as premade building blocks for creating coordination-based hierarchical superstructures. Considering these developments, it may be worth combining the photophysical properties of porphyrin with the shape-persistent porous nature of porphyrin boxes to explore other novel applications. This Account summarizes our recent work on porphyrin boxes, starting with their design, structural features, and applications in different fields. We also try to provide scientific insight into the future opportunities that these amazing boxes have in store for exploring the still uncharted challenging domains in the field of supramolecular chemistry in a confined space.

Structural Control of Metal-Organic Framework Bearing N-Heterocyclic Imidazolium Cation and Generation of Highly Stable Porous Structure.

Building a well-defined porous structure with permanent porosity in a metal-organic framework (MOF) is significant to achieve selective adsorption of guest molecules and its practicality. There have been various efforts to develop new functional frameworks, but chasing both advantages (functions and stability) is always challenging. Herein, we report a highly stable and porous 3D MOF having an imidazolium group. By adjusting counteranions of the imidazolium group, we could prepare two distinct MOFs. The 2D MOF constructed by applying bicarbonate anion inhibited three-dimensional growth and showed N2 adsorption with BET results of 330 m2/g. However, the 3D MOF (Imi-3D) formed by applying chloride anion on the imidazolium group evolved into a 4-fold interpenetrated cube structure. The Imi-3D has high porosity in spite of the folded nature of the structure, with BET results of 1400 m2/g. Interestingly, unlike the usual trend of unstable zinc carboxylate based MOFs, the multiple π-π interactions of the Imi-3D presents a unique tolerance toward various solvents, especially toward water, for 6 months and without decomposition or collapse of the framework. Moreover, since the porous 3D MOF consists of imidazolium groups, the MOF shows a cationic feature with strong adsorption toward anionic guests in aqueous media.

Cucurbit[7]uril-conjugated dyes as live cell imaging probes: investigation on their cellular uptake and excretion pathways.

Here we report the endocytosis and excretion pathways of two different dye-conjugated cucurbit[7]urils, (cyanine 3-conjugated CB[7] and rhodamine X-conjugated CB[7]), which have great potential as molecular probes for live cell imaging. The dye-CB[7]s are translocated into live cells (human breast carcinoma cells, MCF-7) via multiple pathways, predominantly by clathrin-mediated endocytosis, and excreted from cells via lysosome-associated exocytosis. Interestingly, the CB[7] moiety has a substantial influence on the uptake and excretion pathways. These findings may widen the applications of the dyes conjugated to CB[7] and assist in the design of new molecular probes for live cell imaging.

Fuel-Driven Transient Crystallization of a Cucurbit[8]uril-Based Host-Guest Complex.

Transient self-assembling systems often suffer from accumulation of chemical wastes that interfere with the formation of pristine self-assembled products in subsequent cycles. Herein, we report the transient crystallization of a cucurbit[8]uril-based host-guest complex, preventing the accumulation of chemical wastes. Base-catalyzed thermal decarboxylation of trichloroacetic acid that chemically fuels the crystallization process dissolves the crystals, and produces volatile chemical wastes that are spontaneously removed from the solution. With such self-clearance process, no significant damping in the formation of the crystals was observed. The morphology and structural integrity of the crystals was also maintained in subsequent cycles. The concept may be further extended to obtain other temporally functional materials, quasicrystals, etc., based on stimuli-responsive guest molecules.

Rational Design and Construction of Hierarchical Superstructures Using Shape-Persistent Organic Cages: Porphyrin Box-Based Metallosupramolecular Assemblies.

We report a new approach to building hierarchical superstructures using a shape-persistent porous organic cage, which acts as a premade secondary building unit, and coordination chemistry. To illustrate the principle, a zinc-metalated porphyrin box (Zn-PB), a corner-truncated cubic porous cage, was connected by suitable dipyridyl terminated bridging ligands to construct PB-based hierarchical superstructures (PSSs). The PSSs were stabilized not only by the coordination bonds between Zn ions and bipyridyl-terminated ligands but also by π-π interactions between the corners of the Zn-PB units. By varying the length of the linker, we identified an optimum range of the linker length for construction of PSSs. The PSSs have large void volumes and an extrinsic surface area compared to the parent PBs, which can be exploited for the selective encapsulation and interior functionalization of the PSSs for various applications, including catalysis. We observed that singlet oxygen induced synthesis of the natural product, juglone, is more efficiently catalyzed by PSS-1 than its constituent component Zn-PB.

Smart SERS Hot Spots: Single Molecules Can Be Positioned in a Plasmonic Nanojunction Using Host-Guest Chemistry.

Single-molecule surface-enhanced Raman spectroscopy (SERS) offers new opportunities for exploring the complex chemical and biological processes that cannot be easily probed using ensemble techniques. However, the ability to place the single molecule of interest reliably within a hot spot, to enable its analysis at the single-molecule level, remains challenging. Here we describe a novel strategy for locating and securing a single target analyte in a SERS hot spot at a plasmonic nanojunction. The "smart" hot spot was generated by employing a thiol-functionalized cucurbit[6]uril (CB[6]) as a molecular spacer linking a silver nanoparticle to a metal substrate. This approach also permits one to study molecules chemically reluctant to enter the hot spot, by conjugating them to a moiety, such as spermine, that has a high affinity for CB[6]. The hot spot can accommodate at most a few, and often only a single, analyte molecule. Bianalyte experiments revealed that one can reproducibly treat the SERS substrate such that 96% of the hot spots contain a single analyte molecule. Furthermore, by utilizing a series of molecules each consisting of spermine bound to perylene bisimide, a bright SERS molecule, with polymethylene linkers of varying lengths, the SERS intensity as a function of distance from the center of the hot spot could be measured. The SERS enhancement was found to decrease as 1 over the square of the distance from the center of the hot spot, and the single-molecule SERS cross sections were found to increase with AgNP diameter.

Ultrastable Artificial Binding Pairs as a Supramolecular Latching System: A Next Generation Chemical Tool for Proteomics.

In this Commentary, we discuss cucurbit[7]uril-based ultrastable artificial binding pairs as a supramolecular latching system and how we envision this becoming important tools in proteomics. The limitations of current proteomic techniques are described with an emphasis on the lack of tools to answer questions about the complex and dynamic nature of the proteome. Our thoughts as to how artificial ultrastable binding pairs may be able to address these questions are given especially when they are combined with existing methods.