Project Leader: Stefan Matile

  • The general objective of this project is to design, synthesize and use molecules that can 1) move across membranes, and 2) report on their nature.  Considering that the delivery of probes and drugs to their intracellular targets remains a central challenge in chemical biology despite decades of intense research in industry and academia, we felt that conceptual innovation will be unavoidable to achieve significant progress.  Toward this end, we will apply lessons from dynamic covalent chemistry to gain rapid access to libraries of amphiphiles to screen for siRNA uptake, or grow cell-penetrating poly(disulfide)s directly on probes and drugs for their covalent delivery.  Studies toward fluorescent membrane probes focus on conceptual innovation as well.  New families of fluorophores will be designed and synthesized.  Their characteristics will be tested in the broadest sense.  Ultimately, they are expected to sense biologically relevant properties of lipid bilayer membranes including phase, fluidity, homogeneity, microenvironments, tension and potential.

Dynamic amphiphiles

Large collections of dynamic amphiphiles will be generated in situ by the spontaneous reaction of charged hydrophilic “heads” with lipophilic “tails”.  Variables include number and nature of charges (e.g., guanidinium, carboxylate), “heads” (e.g., peptide dendrons), “tails” (e.g., short, long, linear, branched, cyclic, aromatic, spherical, fragrant, fluorophilic, mesogenic, fluorescent, panchromatic), and dynamic covalent bonds (e.g., hydrazones, disulfides, oximes).  Dynamic counterion activation of RNA, DNA and CPP polyions will be identified in fluorogenic large unilamellar vesicles and characterized with regard to applications such as sensing, labeling and delivery. Focused libraries with promising candidates will be screened for siRNA uptake.  Leads from standard RNAi protocols in HeLa cells will be applied to more challenging cell lines used by other members of the NCCR.  Recent publication:  Chem. Eur. J. 2012, 18, 10436.

Cell-penetrating poly(disulfide)s

Lessons from surface-initiated self-organizing polymerization of multicomponent photosystems will be applied to polymerize cell-penetrating poly(disulfide)s directly on substrates right before cellular uptake.  Their depolymerization right after uptake will release unmodified substrates and minimize toxicity.  This approach is expected to afford a general method for the covalent delivery of unmodified probes and drugs.  Initial studies in solution (GPC, molecular weight) and in vesicles (activity in membranes) will be followed by uptake assays (flow cytometry, confocal microscopy imaging) with cell-penetrating poly(disulfide)s grown on fluorescent initiators to systematically optimize their activity (co-polymers; hydrophobicity, halogen bonds, anion-pi interactions, boronic acids, etc).  Optimized cell-penetrating poly(disulfide)s will be applied to deliver probes of interest in other projects of the NCCR.  For surface-initiated self-organizing polymerization, see:  Nature Chem. 2012, 4, 746; halogen bonds:  Nature Commun. 2012, 3, 905. 

New fluorophores

This project focuses on the synthesis of new fluorophores.  Their optoelectronic and functional properties in environments such as bulk and lipid mono-/bilayer membranes, films and surfaces will be characterized.  Topics of interest include FRET systems, fluidity-, homogeneity-, tension- and potential-sensitive membrane probes, panchromatic photosystems as well as supramolecular photovoltaics. 

Triangulenium fluorophores.  The focus of this project is on facile optimization and derivatization of triangulenium fluorophores through ease of synthesis and modularity, and on functional characterization in different environments. As an initial screen we grow giant unilamellar vesicles (GUVs) that show lipid phase partitioning. This allows us to visualize if a probe is preferentially localized into liquid ordered or disordered phases. For probes that do not show partitioning we can examine if the spectral properties change depending upon the lipid environment.  Recent publication:  Dalton Trans. 2012, 41, 6777.

Planarizable push-pull probes.  The objective of this project is to explore whether or not fluorophore planarization and polarization could be combined to obtain conceptually innovative membrane probes.  This mechanism of action applies in many biological processes - reaching from the chemistry of vision to lobster pigmentation – but is poorly explored in the context of fluorescent probes.  Initial studies focus on push-pull oligothiophenes with a partially deplanarized scaffold.  Preliminary results reveal much solvatochromism and thermochromism as well as sensitivity toward membrane fluidity and potentials.  These results validate the concept, provide clear guidelines for the continuation, and promise attractive applications.  With donors and acceptors as good as it gets for now, current efforts focus on the length of push-pull fluorophore and, most importantly, on their twist.  Applications with focus on fluorescent probes for processes of interest in other projects of the NCCR, particularly membrane tension.  Recent publication:  Angew. Chem. Int. Ed. 2012, 51, 12736.

Fluorescent membrane probes

Fluorescent probes will be developed to detect membrane properties such as composition, homogeneity, phase, tension and potential.  New fluorophores such as triangulenium or planarizable push-pull oligothiophenes will be applied as outlined above.  Other topics of interest include probes for liquid-ordered phase based on sphingomyelin mimics, and probes for microenvironments as outlined below.

Exploring the membrane microenvironment surrounding membrane proteins.  The specific objective of this project is the design, synthesis and application of new hybrid probes that can report on the nature of the local environment of membrane proteins. We are using the insulin receptor as a test system. The receptor will be modified with a bifunctional probe: One side of the probe binds covalently to the receptor and the other is a fluorescent sensor that will localize within the lipid bilayer. The distance from receptor can be controlled by varying the length of the linker between the two functional groups.