Rational design of a photoswitchable DNA glue enabling high regulatory function and supramolecular chirality transfer

Short, complementary DNA single strands with mismatched base pairs cannot undergo spontaneous formation of duplex DNA (dsDNA). Mismatch binding ligands (MBLs) can compensate this effect, inducing the formation of the double helix and thereby acting as a molecular glue. Here, we present the rational design of photoswitchable MBLs that allow for reversible dsDNA assembly by light. Careful choice of the azobenzene core structure results in excellent band separation of the E and Z isomers of the involved chromophores. This effect allows for efficient use of light as an external control element for duplex DNA formation and for an in-depth study of the DNA–ligand interaction by UV-Vis, SPR, and CD spectroscopy, revealing a tight mutual interaction and complementarity between the photoswitchable ligand and the mismatched DNA. We also show that the configuration of the switch reversibly dictates the conformation of the DNA strands, while the dsDNA serves as a chiral clamp and translates its chiral information onto the ligand inducing a preference in helical chirality of the Z isomer of the MBLs.

We present the rational design of photoswitchable DNA glue to trigger the reversible formation of duplex DNA by light. The supramolecular assembly shows a mutual interaction between ligand and DNA, which induces a preferred helicity in the switch.     

Allosteric regulation of protein kinase PKCζ by the N-terminal C1 domain and small compounds to the PIF-pocket

Protein kinases are key mediators of cellular signaling, and therefore, their activities are tightly controlled. AGC kinases are regulated by phosphorylation and by N- and C-terminal regions. Here, we studied the molecular mechanism of inhibition of atypical PKCζ and found that the inhibition by the N-terminal region cannot be explained by a simple pseudosubstrate inhibitory mechanism. Notably, we found that the C1 domain allosterically inhibits PKCζ activity and verified an allosteric communication between the PIF-pocket of atypical PKCs and the binding site of the C1 domain. Finally, we developed low-molecular-weight compounds that bind to the PIF-pocket and allosterically inhibit PKCζ activity. This work establishes a central role for the PIF-pocket on the regulation of PKCζ and allows us to envisage development of drugs targeting the PIF-pocket that can either activate or inhibit AGC kinases.     

Time-resolved photoelectron spectroscopy of solvated electrons in aqueous NaI solution

Time-resolved photoelectron spectroscopy was used to study the energetics and dynamics of solvated electrons in aqueous solution. Solvated electrons are generated by ultrafast photodetachment in a 100 mM aqueous NaI solution. Initially, an ensemble of strongly bound ("cold") solvated electrons and an ensemble of weakly bound ("hot") electrons in an unequilibrated solvent environment are observed. We report an ultrafast recombination channel for the "hot" electrons with a rate of (800 fs)(-1) which is in competition with thermalization occurring with a rate of (1.1 ps)(-1). The thermalized electrons recombine with the iodide radical with a rate of (22 ps)(-1). About 35% of the thermalized electrons escape geminate recombination and form free, solvated electrons. The vertical detachment energy for the solvated electron is determined to be 3.40 eV. No indication for a surface-bound electron at lower binding energies was observed.     

Colorimetric quantification of mRNA expression in rare tumour cells amplified by multiple ligation-dependent probe amplification

An enzyme-linked oligonucleotide assay (ELONA) for quantification of mRNA expression of five genes involved in breast cancer, extracted from isolated rare tumour cells and amplified by multiplex ligation-dependent probe amplification (MLPA) is presented. In MLPA, a multiplex oligonucleotide ligation assay is combined with a PCR reaction in which all ligation products are amplified by use of a single primer pair. Biotinylated probes complementary to each of the target sequences were immobilised on the surface of a streptavidin-coated microtitre plate and exposed to single-stranded MLPA products. A universal reporting probe sequence modified with horseradish peroxidase (URP–HRP) and complementary to a universal primer used during the MLPA step was further added to the surface-bound duplex as a reporter probe. Simultaneous addition of anchoring probe and target, followed by addition of reporter probe, rather than sequential addition, was achieved with no significant effect on sensitivity and limits of detection, but considerably reduced the required assay time. Detection limits as low as 20 pmol L⁻¹, with an overall assay time of 95 min could be achieved with negligible cross-reactivity between probes and non-specific targets present in the MLPA-PCR product. The same MLPA-PCR product was analysed using capillary electrophoresis, the technique typically used for analysis of MLPA products, and good correlation was observed. The assay presented is easy to carry out, relatively inexpensive, rapid, does not require sophisticated instrumentation, and enables quantitative analysis, making it very promising for the analysis of MLPA products.     

Optical response of phenantroimidazole-dyes covalently linked to a polynorbornene-backbone to acid and base

Two norbornene derivatives bearing phenantroimidazole dyes, namely endo/exo-11-(4-(1H-phenanthro[9,10-d]imidazol-2-yl)phenoxy) undecylbicyclo[2.2.1] hept-5-ene-2-carboxylat (5) and 11-(4-(1-Methyl-1H-phenanthro[9,10-d]imidazol-2-yl)phenoxy) undecylbicyclo[2.2.1]hept-5-ene-2-carboxylate (6) were synthesized. 5 and 6 were used to prepare statistical copolymers with endo,exo-bicyclo[2.2.1.]hept-2-ene-5,6-dicarboxylic acid dimethylester (7) via ring opening metathesis polymerization. The photophysical properties of the monomers and polymers and to what extent polymerization and substitution patterns influence the photophysical properties were investigated. Furthermore, the optical response upon addition of acid was investigated. The emission of monomer 5 shows a small bathochromic shift of 13 nm upon protonation while the emission of the resultant polymer poly5/7 exhibits a large red shift of 62 nm. In contrast, the according methylated monomer 6 and the resulting polymer poly6/7 gave similar absorption and emission characteristics. The differences were attributed to an increased tendency of the non-methylated dyes to interact when brought in close proximity in the polymer.     

Operating regimes and performance optimization in mode-locked fiber lasers

We derive a theoretical model to characterize the mode-locking dynamics in a single-mode fiber laser cavity with a combination of waveplates and a passive polarizer. The averaging process results in the cubic-quintic Ginzburg-Landau equation (CQGLE) where all the coefficients depend explicitly on the setting of the waveplates as well as the fiber birefringence. A comparison between full numerical simulations and the CQGLE shows a good agreement that allows for characterizing the stability and operating regimes of the laser cavity. A low-dimensional model is developed via the method of proper orthogonal decomposition (POD) to study the multi-pulsing transition of the CQGLE, and the results agree qulitatively with the CQGLE model. The theory allows one to develop guidelines for engineering and optimizing high-energy, high peak-power pulses in the laser cavity.     

General Principles for the Design of Visible-Light-Responsive Photoswitches: Tetra-ortho-Chloro-Azobenzenes

Molecular photoswitches enable reversible external control of biological systems, nanomachines, and smart materials. Their development is driven by the need for low energy (green-red-NIR) light switching, to allow non-invasive operation with deep tissue penetration. The lack of clear design principles for the adaptation and optimization of such systems limits further applications. Here we provide a design rulebook for tetra-ortho-chloroazobenzenes, an emerging class of visible-light-responsive photochromes, by elucidating the role that substituents play in defining their key characteristics: absorption spectra, band overlap, photoswitching efficiencies, and half-lives of the unstable cis isomers. This is achieved through joint photochemical and theoretical analyses of a representative library of molecules featuring substituents of varying electronic nature. A set of guidelines is presented that enables tuning of properties to the desired application through informed photochrome engineering.