Karola Rück-Braun

1,3-Dipolar cycloaddition on solid supports: nitrone approach towards isoxazolidines and isoxazolines and subsequent transformations

1,3-dipolar cycloaddition reactions of nitrones with alkenes and alkynes are well-studied reactions in solution-phase organic chemistry. However, the number of studies concerned with their application in solid-phase organic synthesis is rather low compared to other 1,3-dipoles, e.g. azides or nitrile oxides. This tutorial review aims to summarise the main approaches towards the application of nitrones in 1,3-dipolar cycloaddition reactions on solid supports in addition to subsequent transformations with polymer-bound isoxazolidines and reactions using polymer-bound catalysts.

Photomodulation of ionic current through hemithioindigo-modified gramicidin channels

Incorporation of photo-switchable amino acids into peptides and proteins offers prospects for the control of complex biochemical processes using light. Currently, only a few photo-switchable amino acids are known. We report the design and synthesis of a novel hemithioindigo-based amino acid and its incorporation into the model ion channel gramicidin. Photoisomerization of the hemithioindigo moiety between E and Z isomeric forms is shown to modulate ionic current through the channel in a predictable way. This new amino acid thus expands the possibilities for photo-control in diverse systems.

Reversible photoisomerization of an azobenzene-functionalized self-assembled monolayer probed by sum-frequency generation vibrational spectroscopy

Sum-frequency generation (SFG) vibrational spectroscopy is employed to investigate the reversible, photoinduced trans/cis isomerization of an azobenzene-functionalized self-assembled monolayer (SAM) on a gold substrate. A C[triple bond]N marker group at the outer phenyl ring is used as a direct measure of the switching state. The azobenzene unit is connected to the surface by a tripodal linker system with an adamantane core, which results in both a sufficient decoupling of the functional azobenzene unit from the metallic substrate and a free volume to prevent steric hindrance, thus allowing the isomerization process. Optical excitation at 405 nm induces the trans-->cis isomerization, whereas light exposure at 470 nm leads to the back reaction. The effective cross sections for the reactions are sigma(eff)(cis) = 4 +/- 1 x 10(-18) cm(2) at 405 nm (trans-->cis) and sigma(eff)(trans) = 2.5 +/- 0.9 x 10(-19) cm(2) at 470 nm (cis-->trans). We propose that the photoisomerization is driven by a direct (intramolecular) electronic excitation of the azobenzene conjugate, analogous to the free molecules in solution.

Light‐Directed Protein Binding of a Biologically Relevant β‐Sheet

b-Hairpin structures are frequently involved in protein– protein interactions that control essential processes in cells and are therefore interesting targets for interference. Hairpinforming peptides that compete with such protein interactions are valuable tools for studying biological processes. Moreover, the incorporation of a photoswitchable unit into appropriate b-hairpin-forming peptide ligands could allow protein interactions in cells to be studied by light-triggered interference. However, b-hairpin structures are rarely studied because of the limited availability and stability of suitable model peptides. This is because such a model peptide has to fulfill at least three requirements: 1) the b-hairpin has to be sufficiently stable as monomer without the tendency to selfaggregate, 2) the photoswitchable unit incorporated must stabilize the biologically active peptide conformation, and 3) disturbing the protein binding site by light-induced isomerization of the photoswitch must not result in intermolecular association or even formation of insoluble fibrils. Herein, we report the first example of a b-hairpin model peptide of a biologically important protein domain that shows considerably different binding affinities for the target protein that are dependent on the isomerization state of the embedded photoswitch. PDZ domains mediate the formation of a variety of multiprotein complexes in the cell. Besides C-terminal protein sequences, PDZ domains are also able to recognize internal peptide motifs that bind at the same binding pocket as the C-terminal ones. The best example of this type of internal ligand recognition is found in the extended PDZ domain of neuronal nitric oxide synthase (nNOS) which interacts with the PDZ domain from a-1-syntrophin or the second PDZ domain from PSD95. The formation of the PDZ/PDZ heterodimer requires the b-finger structure of nNOS (30 amino acid residues) to bind at the syntrophin PDZ domain, thus mediating the membrane association of nNOS to skeletal muscle and inducing the production of the second messenger nitric oxide (NO) for muscle contraction. Crucial for binding is the internal recognition motif -LETTFof the extended PDZ domain of nNOS located in the first strand of the hairpin peptide (Scheme 1), a stable

The Hammett Relationship and Reactions in the Excited Electronic State : Hemithioindigo Z/E-Photoisomerization

The photochemical reaction dynamics of a set of photochromic compounds based on thioindigo and stilbene molecular parts (hemithioindigos, HTI) are presented. Photochemical Z/E isomerization around the central double bond occurs with time constants of 216 ps (Z --> E) and 10 ps (E --> Z) for a 5-methyl-hemithioindigo. Chemical substitution on the stilbene moiety causes unusually strong changes in the reaction rate. Electron-donating substituents in the position para to the central double bond (e.g., para-methoxy) strongly accelerate the reaction, while the reaction is drastically slowed by electron-withdrawing groups in this position (e.g., para-nitrile). We correlate the experimental data of seven HTI-compounds in a quantitative manner using the Hammett equation and present a qualitative explanation for the application of ground-state Hammett constants to describe the photoisomerization reaction.

Photocontrol of Contracting Muscle Fibers

The light-controlled inhibition of physiologically relevant protein–protein interactions by appropriate photoresponsive ligands in living cells or small organs (skeletal muscle fibers, vessels) could make it possible to investigate signaling pathways under high spatiotemporal control. Recently, we have reported a cyclic peptide that mimics the b-finger motif in neural NO synthase (nNOS) which is crucial for binding of nNOS to a-1-syntrophin. When a photoswitchable unit is embedded into this peptide, the binding can be controlled in vitro simply by light. In skeletal muscle the extended PDZ domain of nNOS interacts with the PDZ domain of a-1syntrophin to recruit nNOS to the dystrophin-associated protein complex in the plasma membrane, thus coupling the production of the second messenger nitric oxide (NO) to muscle contraction. Loss of sarcolemmal nNOS is known to result in functional ischemia during muscle contraction, which is commonly observed in muscle diseases as Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD). Herein we show that the photoswitchable peptide ligand 1 (Figure 1) is able to translocate into cells, is sufficiently stable towards intracellular conditions, and can be used in vivo to photocontrol contracting muscle fibers. The recently described nNOS-derived, photoswitchable peptide ligand 1 of a-1-syntrophin contains the azobenzenew-amino acid 3-((4’-aminomethyl)phenylazo)benzoic acid (3,4’-AMPB), which in its trans form led to a ligand that showed no affinity to the PDZ domain of a-1-syntrophin while photoisomerization to the cis form resulted in a remarkable affinity of the peptide (KD = 10.6 mm). [1a] The finding has addressed the question of the applicability of the light-controlled ligand under physiological conditions to investigate the native interference in living skeletal muscles. Light-directed binding of the cis form of the photoswitchable ligand 1 to syntrophin was expected to inhibit the native syntrophin–nNOS interaction in the skeletal muscle followed by the dislocation of nNOS from the sarcolemma which may result in reduced NO release from skeletal muscle cells and thus in light-controlled muscle contraction. Azobenzene units have been used extensively for the photomodulation of biomolecules (peptides, proteins, and nucleic acids) and biological processes in vitro and in vivo (as in ion channels). The feasibility of using azobenzene systems in living organisms to photocontrol biological events has been confirmed by the in vivo imaging of the isomerization process in zebrafish. An intrinsic hindrance for applications of azobenzene in living cells derives from its susceptibility to reduction. The azo unit may be subject to reduction by enzymes or thiols such as glutathione (GSH) which is present in most cells at millimolar concentration (0.5–10 mm). The reduction rate of the cis isomer of a parasubstituted AMPB amino acid in a model tripeptide is about 100-fold higher than for the corresponding trans isomer. To determine the stability of the AMPB switch unit in the peptide ligand 1 we incubated the cis form of the photoswitchable ligand at the photostationary state (pss) in buffer solution (pH 7.5) containing reduced glutathione (10 mm). After 1 h exposure to GSH no reduced material was detectable by LC–MS analysis (Figure 2, dashed line). Even after 16 h the reduced material amounted to only 5% (Figure 2, dotted line, signal marked with an arrow; Figure S3 in the Supporting Information). In addition, when changes in the UV/Vis spectra of the photoswitchable ligand were followed during irradiation in buffer solution (pH 7.5) containing GSH (10 mm), isosbestic points were retained, indicating the stability of the 3,4’-AMPB unit in the peptide ligand 1 (Figure S1 in the Supporting Information). As expected, the thermal cis!trans isomerization of the photoswitchable ligand 1 in the presence of 10 mm glutathione was Figure 1. Structure of the cis form of the photoswitchable peptide ligand related to the b-finger peptide of nNOS.

Light-Switchable Hemithioindigo-Hemistilbene-Containing Peptides : Ultrafast Spectroscopy of the Z -> E Isomerization of the Chromophore and the Structural Dynamics of the Peptide Moiety

Two hemithioindigo-hemistilbene (HTI) derivatives, designed to operate as structural switches in peptides, as well as two HTI peptides are characterized by ultrafast spectroscopy in the visible and the infrared. The two HTI switches follow the reaction scheme published for other HTI compounds with a picosecond excited state reaction (τ(1) ≈ 6 ps) and isomerization from Z to E with τ(2) = 13 and 51 ps. As compared to the isolated chromophores, the isomerization reaction is slowed down in the chromopeptides to τ(2) = 24 and 69 ps. For the smaller peptide containing 6 amino acids, the structural changes of the peptide moiety observed via the IR spectrum in the amide I band follow the isomerization of the molecular switch closely. In the larger cyclic chromopeptide, containing 20 amino acids and mimicking a β-hairpin structure in the Z-form of the chromophore, the peptide moiety also changes its structure during isomerization of the chromophore. However, the IR spectrum at the end of the observation period of 3 ns deviates significantly from the stationary difference spectrum. These signatures indicate that strong additional structural changes, e.g., breaking of interchain hydrogen bonds, also occur on longer time scales.

Chiral auxiliaries in cycloadditions

[2+1] Cycloadditions (Cyclopropanations). [2+2] Cycloadditions. [4+2] Cycloadditions (Diels-Alder Reactions). Hetero [4+2] Cycloadditions. [3+2]Cycloadditions. Chiral Auxiliaries. Subject Index.

Photoactivation of an Inhibitor of the 12/15-Lipoxygenase Pathway

Lipoxygenases are lipid‐peroxidizing enzymes that have been implicated in the pathogenesis of inflammatory diseases and lipoxygenase inhibitors may be developed as anti‐inflammatory drugs. Structure comparison with known lipoxygenase inhibitors has suggested that (2Z)‐2‐(3‐benzylidene)‐3‐oxo‐2,3‐dihydrobenzo[b]thiophene‐7‐carboxylic acid methyl ester might inhibit the lipoxygenase pathway but we found that it exhibited only a low inhibitory potency for the pure 12/15‐lipoxygenase (IC50=0.7 mM). However, photoactivation, which induces a Z‐to‐E isomerization of the double bond, strongly augmented the inhibitory potency and an IC50 value of 0.021 mM was determined for the pure E isomer. Similar isomer‐specific differences were observed with the recombinant enzyme and its 12‐lipoxygenating Ile418Ala mutant, as well as in intracellular lipoxygenase activity. Structure modeling of the enzyme/inhibitor complex suggested the molecular reasons for this isomer specificity. Since light‐induced isomerization may proceed in the skin, such photoreactive compounds might be developed as potential drugs for inflammatory skin diseases.

Light-Switchable Peptides with a Hemithioindigo Unit : Peptide Design, Photochromism, and Optical Spectroscopy

This Minireview focuses on the hemithioindigo photoswitch and its use for the reversible control of three-dimensional peptide structure and related biological functions. Both the general design aspects and biophysical properties of various hemithioindigo-based chromopeptides are summarized. Hemithioindigo undergoes reversible Z→E photoisomerization after absorption of visible light. The unique ultrafast switching mechanism of hemithioindigo combines picosecond isomerization kinetics with strong double-bond torsion after light absorption, making it the ideal tool for instantaneous modulation of biological structure. Various inhibitors and model peptides based on hemithioindigo are described that can directly regulate biological signaling or allow the fastest events in peptide folding to be studied. Finally, a diverse range of chromopeptides with photoswitchable β-hairpin structures based on azobenzenes, stilbenes, and hemithioindigo are compared to emphasize the unique properties of hemithioindigo.