Membrane fusion of vesicles of oligomerisable lipids

his thesis aimed to support the current model of membrane fusion by (1) providing an avenue towards ultrastructural characterisation of hypothetical membrane fusion intermediates, such as the stalk(2) extending the model to calcium-induced membrane funsion and (3) analysing the thermodynamic characteristics of membrane fusion. ... Zie: Summar

Orthogonal, metal-free surface modification by strain-promoted azide–alkyne and nitrile oxide–alkene/alkyne cycloadditions

In this article we present a fast and efficient methodology for biochemical surface patterning under extremely mild conditions. Micropatterned azide/benzaldoxime-surfaces were prepared by microcontact printing of a heterobifunctional cyclooctyne oxime linker on azide-terminated self-assembled monolayers (SAMs). Strain-promoted azide–alkyne cycloaddition (SPAAC) in combination with microcontact printing allows fast and effective surface patterning. The resulting bifunctional azide/oxime substrates could successfully be used for metal-free, orthogonal immobilization of various biomolecules by 1,3-dipolar cycloadditions at room temperature. Azide-decorated areas were modified by reaction with a cyclooctyne-conjugate using SPAAC, while benzaldoxime-decorated areas were activated by in situ oxidation to the reactive nitrile oxides and subsequent nitrile oxide cycloaddition with alkene- and alkyne-functionalized bioconjugates. In addition, orthogonal double immobilization was achieved by consecutive and independent SPAAC and nitrile oxide cycloadditions

Metal ion responsive adhesion of vesicles by conformational switching of a non-covalent linker

This contribution describes the metal ion responsive adhesion of vesicles induced by a conformational switch of a non-covalent linker molecule. A p-tert-butylbenzyl dimer with a flexible N,N'-bis(3-aminopropyl)ethylenediamine spacer was used as a non-covalent linker, which induces aggregation and adhesion (but not fusion) of host bilayer vesicles composed of amphiphilic beta-cyclodextrins by the formation of hydrophobic inclusion complexes. The aggregation and adhesion of the vesicles in dilute aqueous solution was confirmed by isothermal titration calorimetry (ITC), optical density measurements at 600 nm (OD600), dynamic light scattering (DLS), zeta-potential measurements, cryogenic transmission electron microscopy (cryo-TEM) and fluorescence spectroscopy. However, in the presence of a divalent metal ion like Cu(2+), the tetra-amine linker molecule forms a stable metal coordination complex and dramatically switches its conformation from linear to bent, which results in the dissociation of intervesicular complexes, and leads to the dispersion of vesicle clusters. This process is reversible in the presence of a strong metal ion chelator, such as EDTA, that scavenges the Cu(2+) ion complexed by the linker. The linker molecule regains its linear conformation and triggers the reaggregation of the vesicles. In contrast, conformational switching was inhibited by introducing a rigid N,N'-bis(3-aminopropyl)piperazine spacer in the non-covalent linker molecule and vesicles do not aggregate in the presence of a cyclic guest that can only bind intravesicularly. Thus, a metal ion regulated molecular switch can control the aggregation state of an organic colloidal solution.</p

Solid-supported monolayers and bilayers of amphiphilic ß-Cyclodextrin

This paper describes the adsorption and spreading of B-cyclodextrin (CD) vesicles on hydrophobic and hydrophilic substrates, which involves a transition from bilayer vesicles to planar molecular monolayers or bilayers. On substrates that are patterned with self-assembled monolayers by microcontact printing (..CP), the CD vesicles preferentially adsorb on hydrophobic areas instead of hydrophilic (nonionic) areas, and on cationic areas instead of hydrophilic (nonionic) areas. Supported monolayers of amphiphilic cyclodextrins CD1 and CD2 were obtained by adsorption of CD vesicles to hydrophobic substrates, and supported bilayers of amphiphilic cyclodextrins CD1 and CD2 were prepared by adsorption of CD vesicles on cationic substrates. Contact angle goniometry, atomic force microscopy and confocal fluorescence microscopy (CFM) were used to analyze the supported CD layers. The fluidity of the supported CD layers was verified using fluorescence recovery after photobleaching experiments. The supported layers function as a supramolecular platform that can bind suitable guest molecules through inclusion in the CD host cavities. Additionally, the CD host layers were patterned with fluorescent guest molecules by supramolecular ..CP on the supported CD layers. The host-guest interactions were investigated with CFM and fluorescence resonance energy transfer experiments

A carbohydrate-based hydrogel containing vesicles as responsive non-covalent cross-linkers

In this edge article we report the preparation of a supramolecular carbohydrate hydrogel containing cyclodextrin vesicles as 3D junctions. A cellulose polymer is randomly modified with hydrophobic side groups that act as guests for the cyclodextrin hosts on the surface of the vesicles. Hence, the vesicles interconnect the polymer chains into a three-dimensional network and act as multivalent linkages. The resulting gel shows significant shear-thinning and self-healing properties, which make it highly suitable for applications that require injectability. Furthermore, SAXS and cryo-TEM measurements indicate that intact vesicles are present in the gel matrix

Surface functionalization with carboxylic acids by photochemical microcontact printing and tetrazole chemistry

In this paper, we show that carboxylic acid-functionalized molecules can be patterned by photochemical microcontact printing on tetrazole-terminated self-assembled monolayers. Upon irradiation, tetrazoles eliminate nitrogen to form highly reactive nitrile imines, which can be ligated with several different nucleophiles, carboxylic acids being the most reactive. As a proof of concept, we immobilized trifluoroacetic acid to monitor the reaction with X-ray photoelectron spectroscopy. Moreover, we also immobilized peptides and fabricated carbohydrate−lectin as well as biotin−streptavidin microarrays using this method. Surface-patterning was demonstrated by fluorescence microscopy and time-of-flight secondary ion mass spectrometr

Sequence-selective detection of double-stranded DNA sequences using pyrrole-imidazole polyamide microarrays

We describe a microarray format that can detect double-stranded DNA sequences with a high degree of sequence selectivity. Cyclooctyne-derivatized pyrrole-imidazole polyamides were immobilized on azide-modified glass substrates using microcontact printing and a strain-promoted azide-alkyne cycloaddition (SPAAC) reaction. These polyamide-immobilized substrates selectively detected a seven-base-pair binding site incorporated within a double-stranded oligodeoxyribonucleotide sequence even in the presence of an excess of a sequence with a single-base-pair mismatc