Most azobenzene-based photoswitches use UV light for photoisomerization. This can limit their application in biological systems, where UV light can trigger unwanted responses, including cellular apoptosis. We have found that substitution of all four ortho positions with methoxy groups in an amidoazobenzene derivative leads to a substantial (~35 nm) red shift of the n-π* band of the trans isomer, separating it from the cis n-π* transition. This red shift makes trans-to-cis photoswitching possible using green light (530-560 nm). The cis state is thermally stable with a half-life of ~2.4 days in the dark in aqueous solution. Reverse (cis-to-trans) photoswitching can be accomplished with blue light (460 nm), so bidirectional photoswitching between thermally stable isomers is possible without using UV light at all.

The photoisomerization of azobenzenes provides a general means for the photocontrol of molecular structure and function. For applications in vivo, however, the wavelength of irradiation required for trans-to-cis isomerization of azobenzenes is critical since UV and most visible wavelengths are strongly scattered by cells and tissues. We report here that azobenzene compounds in which all four positions ortho to the azo group are substituted with bulky electron-rich substituents can be effectively isomerized with red light (630-660 nm), a wavelength range that is orders of magnitude more penetrating through tissue than other parts of the visible spectrum. When the ortho substituent is chloro, the compounds also exhibit stability to reduction by glutathione, enabling their use in intracellular environments in vivo.

Biological tissue exhibits an absorbance minimum in the near-infrared between 700 and 900 nm that permits deep penetration of light. Molecules that undergo photoisomerization in this bio-optical window are highly desirable as core structures for the development of photopharmaceuticals and as components of chemical-biological tools. We report the systematic design, synthesis, and testing of an azobenzene derivative tailored to undergo single-photon photoswitching with near-infrared light under physiological conditions. A fused dioxane ring and a methoxy substituent were used to place oxygen atoms in all four ortho positions, as well as two meta positions, relative to the azobenzene N═N double bond. This substitution pattern, together with a para pyrrolidine group, raises the pK of the molecule so that it is protonated at physiological pH and absorbs at wavelengths >700 nm. This azobenzene derivative, termed DOM-azo, is stable for months in neutral aqueous solutions, undergoes trans-to-cis photoswitching with 720 nm light, and thermally reverts to the stable trans isomer with a half-life near 1 s.

Longer switching wavelengths and good photochemical yields and stabilities of the cis isomers in reducing aqueous environments are achieved by introducing 2,2'-aminoalkyl substituents into 4,4'-diamido-substituted azobenzenes. The products are thus suitable for photocontrol of biomolecular structures in intracellular environments, such as switching between two peptide configurations (see picture).

Carbon-11 labeled isocyanates are efficiently prepared by dehydration of [(11) C]carbamate salts, which in turn are easily formed from cyclotron-produced [(11) C]CO(2) and amines in the presence of a CO(2) fixation agent. The [(11) C]isocyanates are useful radiosynthons for the synthesis of a variety of [carbonyl-(11) C]-labeled asymmetrical ureas and carbamate esters. The method is well suited to incorporate any isotope of carbon, and is especially useful for positron emission tomography (PET) radiotracers for in vivo imaging. This is demonstrated by using the method to make [carbonyl-(11) C]-6-hydroxy-[1,1'-biphenyl]-3-yl cyclohexylcarbamate which is a novel radiotracer for PET imaging of fatty acid amide hydrolase.

The functions of peptides and proteins often depend critically on their folded three-dimensional structures. However, the folded states of most proteins are only e10 kcal/mol lower in energy than their unfolded states. 1 Short peptide sequences are usually unfolded in aqueous solution, although they adopt specific folded structures when bound to their biological targets. Stabilizing the active folded forms of peptide or proteins is thus important for maintaining or enhancing the functions of these molecules under a wide variety of conditions. Owing to its prevalence as a secondary structural element in proteins, the R-helix has received considerable attention as a target for conformational stabilization. Stabilized helices have been employed as biological agents, for example, against microbial infections, 2 and for targeting tumor suppressor proteins 3 and proteins involved in apoptosis. 4 Methods developed to stabilize peptide R-helical structures include introduction of R,R-dialkyl amino acids, 5 cross-linking amino acid side chains via disulfide bonds, 6 metal chelates, 7 lactam bridges, 8,9 or via ruthenium-catalyzed ring closing metathesis (RCM) to form cyclic peptides. 10 These methods require peptide synthesis using non-natural amino acids. Recently, Fujimoto et al. introduced acetylenic cross-linking agents that could be introduced via pairs of naturally occurring lysine side chains; 11 however, the degree of helix stabilization was moderate.In the absence of specific favorable interactions between a crosslinker and the folded state of a peptide or protein, the mechanism of stabilization appears to be primarily the decrease in conformational entropy of the unfolded state caused by an intramolecular covalent linkage. 12,13 We anticipate that maximal stabilization should occur with (i) optimal matching between the cross-linker length and the distance distribution between the two attachment points of the folded peptide or protein, (ii) enhanced rigidity of the crosslinker, and (iii) a long cross-linker that can bridge sites far apart in amino acid sequence. Here we introduce a water-soluble, thiolreactive cross-linking reagent (EY-CBS) (3,3′-ethyne-1,2-diylbis-{6-[(chloroacetyl)amino]benzenesulfonic acid} (1) that can be used to introduce long, rigid bridges into peptides and proteins. We compare its stabilizing effects on helices as well as on a small -sheet protein with the effects of more flexible linkers, including the commercially available 1,4-di[3′-(2′-pyridyldithio]propionamido]butane (DPDPB). Figure 1 shows the structure of EY-CBS (1), synthesized in four steps (see Supporting Information), together with an analogue EA-CBS (2) with a single bond in place of the central triple bond, and the flexible commercial cross-linker DPDPB (3). EY-CBS is designed as linear, symmetric, and Cys-reactive to minimize the number of possible conformations of the cross-link with respect to the peptide backbone attachment points. Sulfonate groups are included for water solubility since introduction of a large hydrophob...

Azobenzene derivatives can be used to reversibly photoregulate secondary structure when introduced as intramolecular bridges in peptides and proteins. Here we report the design, synthesis, and characterization of a disubstituted N,N-dialkyl azobenzene derivative that absorbs near 480 nm in aqueous solution and relaxes with a half-life of approximately 50 ms at room temperature. The wavelength of maximum absorbance and the rate of thermal relaxation are solvent-dependent. An increase in the percentage of organic solvent leads, in general, to a blue shift in the absorbance maximum and a slowing of the relaxation rate. In accordance with the design, the thermal relaxation of the azobenzene cross-linker from cis to trans causes an increase in the helix content of one peptide where the linker is attached via cysteine residues spaced at i, i + 11 positions and a decrease in helix content of another peptide with cysteine residues spaced at i, i + 7. This cross-linker design thus expands the possibilities for fast photocontrol of peptide and protein structure.

Längere Schaltwellenlängen und gute photochemische Ausbeuten und Stabilitäten des cis‐Isomers in reduzierenden wässrigen Umgebungen kennzeichnen 4,4′‐Diamido‐substituierte Azobenzole mit 2,2′‐Aminoalkyl‐Substituenten. Die Produkte eignen sich zur Photosteuerung biomolekularer Strukturen in zellulären Umgebungen, etwa zum Umschalten zwischen zwei Peptidkonfigurationen (siehe Bild).