Jens Voskuhl

Light-responsive capture and release of DNA in a ternary supramolecular complex.

The wavelength determines whether DNA is captured in a light-responsive ternary supramolecular complex or released (see scheme). The reversible binding of DNA is triggered by a photoisomerization, which switches the complex from a multivalent to a monovalent binding mode.

Sugar-Decorated Sugar Vesicles: Lectin-Carbohydrate Recognition at the Surface of Cyclodextrin Vesicles

An artificial glycocalix self-assembles when unilamellar bilayer vesicles of amphiphilic beta-cyclodextrins are decorated with maltose and lactose by host-guest interactions. To this end, maltose and lactose were conjugated with adamantane through a tetra(ethyleneglycol) spacer. Both carbohydrate-adamantane conjugates strongly bind to beta-cyclodextrin (K(a) approximately 4 x 10(4) M(-1)). The maltose-decorated vesicles readily agglutinate (aggregate) in the presence of the lectin concanavalin A, whereas the lactose-decorated vesicles agglutinate in the presence of peanut agglutinin. The orthogonal multivalent interaction in the ternary system of host vesicles, guest carbohydrates, and lectins was investigated by using isothermal titration calorimetry, dynamic light scattering, UV/Vis spectroscopy, and cryogenic transmission electron microscopy. It was shown that agglutination is reversible, and the noncovalent interaction can be suppressed and eliminated by the addition of competitive inhibitors, such as D-glucose or beta-cyclodextrin. Also, it was shown that agglutination depends on the surface coverage of carbohydrates on the vesicles.

Dual stimuli-responsive self-assembled supramolecular nanoparticles

Supramolecular nanoparticles (SNPs) encompass multiple copies of different building blocks brought together by specific noncovalent interactions. The inherently multivalent nature of these systems allows control of their size as well as their assembly and disassembly, thus promising potential as biomedical delivery vehicles. Here, dual responsive SNPs have been based on the ternary host-guest complexation between cucurbit[8]uril (CB[8]), a methyl viologen (MV) polymer, and mono- and multivalent azobenzene (Azo) functionalized molecules. UV switching of the Azo groups led to fast disruption of the ternary complexes, but to a relatively slow disintegration of the SNPs. Alternating UV and Vis photoisomerization of the Azo groups led to fully reversible SNP disassembly and reassembly. SNPs were only formed with the Azo moieties in the trans and the MV units in the oxidized states, respectively, thus constituting a supramolecular AND logic gate.

In Situ Modification of Plain Liposomes with Lipidated Coiled Coil Forming Peptides Induces Membrane Fusion

Complementary coiled coil forming lipidated peptides embedded in liposomal membranes are able to induce rapid, controlled, and targeted membrane fusion. Traditionally, such fusogenic liposomes are prepared by mixing lipids and lipidated peptides in organic solvent (e.g., chloroform). Here we prepared fusogenic liposomes in situ, i.e., by addition of a lipidated peptide solution to plain liposomes. As the lipid anchor is vital for the correct insertion of lipidated peptides into liposomal membranes, a small library of lipidated coiled coil forming peptides was designed in which the lipid structure was varied. The fusogenicity was screened using lipid and content mixing assays showing that cholesterol modified coiled coil peptides induced the most efficient fusion of membranes. Importantly, both lipid and content mixing experiments demonstrated that the in situ modification of plain liposomes with the cholesterol modified peptides yielded highly fusogenic liposomes. This work shows that existing membranes can be activated with lipidated coiled coil forming peptides, which might lead to highly potent applications such as the fusion of liposomes with cells.

Molecular Recognition of Vesicles: Host–Guest Interactions Combined with Specific Dimerization of Zwitterions

The aggregation of beta-cyclodextrin vesicles can be induced by an adamantyl-substituted zwitterionic guanidiniocarbonylpyrrole carboxylate guest molecule (1). Upon addition of 1 to the cyclodextrin vesicles at neutral pH, the vesicles aggregate (but do not fuse), as shown by using UV/Vis and fluorescence spectroscopy, dynamic light scattering, zeta-potential measurements, cryogenic transmission electron microscopy, and atomic force microscopy. Aggregation of the vesicles is induced by a twofold supramolecular interaction. First, the adamantyl group of 1 forms an inclusion complex with beta-cyclodextrin. Second, at neutral pH the guanidiniocarbonylpyrrole carboxylate zwitterion dimerizes through the formation of hydrogen-bonded ion pairs. Because the dimerization of 1 depends on the zwitterionic protonation state of 1, the aggregation of the cyclodextrin vesicles is also pH dependent; the cyclodextrin vesicles do not interact at pH 5 or 9, at which 1 is either cationic or anionic and, therefore, not self-complementary. These observations are consistent with molecular recognition of the vesicles through a combination of two different supramolecular interactions, that is, host-guest inclusion and dimerization of zwitterions, at the bilayer membrane surface.

Multivalent Interaction of Cyclodextrin Vesicles, Carbohydrate Guests, and Lectins: A Kinetic Investigation†

An artificial glycocalix self-assembles when unilamellar bilayer vesicles of amphiphilic β-cyclodextrins are decorated with maltose- and lactose-adamantane conjugates by host-guest interactions. The maltose-decorated vesicles aggregate in the presence of lectin concanavalin A whereas the lactose-decorated vesicles aggregate in the presence of lectin peanut agglutinin. The kinetics of the orthogonal multivalent interfacial interactions present in this ternary system of vesicles, carbohydrates, and lectins were studied by time-dependent measurements of the optical density at 400 nm. The average vesicle and vesicle aggregate sizes were monitored by dynamic light scattering. The aggregation process was evaluated as a function of lectin concentration, vesicle concentration, and surface coverage of the vesicles by the carbohydrate-adamantane conjugates. The initial rate of vesicle aggregation scales linearly with the lectin as well as the cyclodextrin vesicle concentration. Furthermore, each lectin requires a characteristic critical density of carbohydrates at the vesicle surface. These observations allow a prediction of the response of the ternary supramolecular system at different concentrations of its components. Also, the effective binding site separation in a multivalent receptor such as a multiple binding site protein can be accurately determined. This methodology can be extended to multivalent noncovalent interactions in other ligand-receptor systems at interfaces.

Advances in contact printing technologies of carbohydrate, peptide and protein arrays

Microcontact printing (μCP) techniques are powerful tools to print molecules on reactive surfaces in a covalent or non-covalent manner to produce well-defined patterns, in shape and spot morphology, of bioactive molecules such as carbohydrates, peptides and proteins. These printed biofunctional surfaces have nowadays found increased use in a range of bioanalytical and biomedical applications, for example, in the investigation of eukaryotic cell and bacteria behavior on solid supports. This review focuses on advances in techniques of μCP over the past three years and some recent appealing applications of the printed arrays are illustrated.

In Vitro and In Vivo Supramolecular Modification of Biomembranes Using a Lipidated Coiled‐Coil Motif

The molecular building blocks available in biological systems self-assemble into defined structures in an extremely controlled manner. These structures must be flexible and adaptive to the environment in order to carry out their function in a regulated manner. Therefore, nature uses multiple weak interactions (e.g. hydrogen bonding and van der Waals interactions) to act as the glue to hold these structures together. When many weak interactions cooperatively combine, relatively stable entities are produced, which retain the ability to respond to external stimuli such as fluctuations in ion concentration, pH, and temperature. For many years, nature has been a source of inspiration for supramolecular chemistry. Scientists typically follow the bottom-up approach and design relatively simple molecules which assemble into functional materials with well-defined properties. Recent progress has resulted in molecular systems that are responsive to multiple stimuli and are therefore highly controlled, emulating nature ever more closely. A relatively new development is the application of supramolecular constructs in in vitro and in vivo environments to directly study and influence biological processes in live cells. Chemically tailored systems can be integrated into cell membranes, for example. This enables the modification or regulation of cellular behavior through external artificial signals. There are two approaches for introducing chemical species into a cell membrane by supramolecular chemistry: 1) specific binding of guest molecules to membrane-anchored biomolecules such as native proteins and 2) nonspecific labeling of membranes with the aid of hydrophobic and electrostatic interactions or through a chemical crosslinker. Lipidated peptides are particularly good candidates for application in biological systems as their aggregation behavior can be controlled by carefully balancing the hydrophobicity of the anchor and the hydrophilicity of the cargo; this aids the incorporation of lipidated peptides into membranes. Here we describe the use of a coiled-coil motif as the peptide segment, a highly specific recognition system that can be introduced into live cells. The coiled-coil motif acts as molecular Velcro and can thus be used to link distinct molecular constructs. An example of the specific labeling of proteins through coiled-coil formation was recently supplied by Matsuzaki et al. Surface modification through the nonspecific binding of polymers to cell membranes has also been studied, for example by Ijiro et al. Lipid-grafted polymers adhere to cell membranes and could potentially act as a scaffold to which a wide range of functional moieties could be attached, thereby intervening in the chemistry of the cell. Furthermore, cationic graft copolymers have also been shown to interact electrostatically with cell membranes, resulting in chemically altered cell membranes. Although these examples illustrate that in vitro membrane functionalization is a highly rewarding strategy, there are currently no examples of efficient in vivo strategies. Therefore, it is our goal to transiently modify lipid membranes through specific supramolecular interactions in in vitro and in vivo environments. For this purpose, we use a pair of complementary coiled-coil-forming lipidated peptides (E and K peptides) to specifically introduce a noncovalent and bio-orthogonal recognition motif to biological membranes (Scheme 1). Here, we describe a generic supramolecular tool which allows us to rapidly and efficiently form coiled-coil motifs at the surface of biological membranes. This is of interest as a wide range of molecular constructs can be introduced to the surface of the cell in this way. Coiled-coil-forming peptides E [(EIAALEK)3] and K [(KIAALKE)3] [12] were first covalently conjugated to PEG12 spacers (PEG= polyethylene glycol). Subsequently, a cholesterol moiety was coupled to the pegylated peptides yielding CPE and CPK (Scheme 1A). The cholesterol moiety allows for the immediate insertion of the lipidated peptides into membranes through hydrophobic interactions and the PEG12 moiety was incorporated to aid efficient molecular recognition between the peptide segments E and K. Recently we showed that upon the addition of micellar solutions of either CPE or CPK to plain liposomes, the lipidated peptides spontaneously inserted into liposomal membranes. In the current study, CHO cell membranes (CHO=Chinese hamster ovary) and the skin of zebrafish embryos were modified with coiled-coil-forming peptides by the addition of a micellar solution of CPE or CPK, resulting in immediate incorporation of these amphiphiles into the membranes. Subsequently, the complementary peptide was added, result[*] M. Sc. H. R. Zope, M. Sc. F. Versluis, Dr. J. Voskuhl, Dr. A. Kros Soft Matter Chemistry, Leiden Institute of Chemistry Leiden University P.O. Box 9502, 2300 RA Leiden (The Netherlands) E-mail: a.kros@chem.leidenuniv.nl

A soft supramolecular carrier with enhanced singlet oxygen photosensitizing properties

Herein we report the self-assembly of a supramolecular singlet oxygen photosensitizing system from an adamantane-functionalized, hexaanionic water-soluble zinc(II) phthalocyanine (PC) and β-cyclodextrin vesicles (CDV). Characterisation of the designed PC, which was synthesized by an asymmetric statistical condensation, was carried out by several analytical techniques such as MALDI-HRMS, NMR, IR, UV/vis as well as steady state and time resolved fluorescence spectroscopy. The influence of the docking of the PC to the CDVs on the PC photoluminescence as well as on the singlet oxygen photoproduction quantum yields was investigated. The results indicate that the host–guest interaction of the photosensitizer and the CDVs significantly prevents the formation of inactive aggregates, and enhances the photosensitizing ability of the PC. The supramolecular assembly constitutes a biocompatible photoactive platform for the design of phototherapeutic agents.

Light‐Triggered Capture and Release of DNA and Proteins by Host–Guest Binding and Electrostatic Interaction

The development of an effective and general delivery method that can be applied to a large variety of structurally diverse biomolecules remains a bottleneck in modern drug therapy. Herein, we present a supramolecular system for the dynamic trapping and light-stimulated release of both DNA and proteins. Self-assembled ternary complexes act as nanoscale carriers, comprising vesicles of amphiphilic cyclodextrin, the target biomolecules and linker molecules with an azobenzene unit and a charged functionality. The non-covalent linker binds to the cyclodextrin by host-guest complexation with the azobenzene. Proteins or DNA are then bound to the functionalized vesicles through multivalent electrostatic attraction. The photoresponse of the host-guest complex allows a light-induced switch from the multivalent state that can bind the biomolecules to the low-affinity state of the free linker, thereby providing external control over the cargo release. The major advantage of this delivery approach is the wide variety of targets that can be addressed by multivalent electrostatic interaction, which we demonstrate on four types of DNA and six different proteins.