Two ground-state modifications of quantum-dot beryllium

Exact electronic properties of a system of four Coulomb-interacting two-dimensional electrons in a parabolic confinement are reported. We show that degenerate ground states of this system are characterized by qualitatively different internal electron-electron correlations, and that the formation of Wigner molecule in the strong-interaction regime is going on in essentially different ways in these ground states.Comment: 5 pages, incl 5 Figures and 2 Table

The two electron artificial molecule

Exact results for the classical and quantum system of two vertically coupled two-dimensional single electron quantum dots are obtained as a function of the interatomic distance (d) and with perpendicular magnetic field. The classical system exhibits a second order structural transition as a function of d which is smeared out and shifted to lower d values in the quantum case. The spin-singlet - spin-triplet oscillations are shifted to larger magnetic fields with increasing d and are quenched for a sufficiently large interatomic distance.Comment: 4 pages, 4 ps figure

Differential Scanning Fluorimetry provides high throughput data on silk protein transitions

Here we present a set of measurements using Differential Scanning Fluorimetry (DSF) as an inexpensive, high throughput screening method to investigate the folding of silk protein molecules as they abandon their first native melt conformation, dehydrate and denature into their final solid filament conformation. Our first data and analyses comparing silks from spiders, mulberry and wild silkworms as well as reconstituted ‘silk’ fibroin show that DSF can provide valuable insights into details of silk denaturation processes that might be active during spinning. We conclude that this technique and technology offers a powerful and novel tool to analyse silk protein transitions in detail by allowing many changes to the silk solutions to be tested rapidly with microliter scale sample sizes. Such transition mechanisms will lead to important generic insights into the folding patterns not only of silks but also of other fibrous protein (bio)polymers

Extrinsic Fluorescent Dyes as Tools for Protein Characterization

Noncovalent, extrinsic fluorescent dyes are applied in various fields of protein analysis, e.g. to characterize folding intermediates, measure surface hydrophobicity, and detect aggregation or fibrillation. The main underlying mechanisms, which explain the fluorescence properties of many extrinsic dyes, are solvent relaxation processes and (twisted) intramolecular charge transfer reactions, which are affected by the environment and by interactions of the dyes with proteins. In recent time, the use of extrinsic fluorescent dyes such as ANS, Bis-ANS, Nile Red, Thioflavin T and others has increased, because of their versatility, sensitivity and suitability for high-throughput screening. The intention of this review is to give an overview of available extrinsic dyes, explain their spectral properties, and show illustrative examples of their various applications in protein characterization

Introduction of Intrinsic Kinetics of Protein–Ligand Interactions and Their Implications for Drug Design

Structure–kinetic relationship analyses and identification of dominating interactions for optimization of lead compounds should ideally be based on <i>intrinsic</i> rate constants instead of the more easily accessible <i>observed</i> kinetic constants, which also account for binding-linked reactions. The intrinsic rate constants for sulfonamide inhibitors and pharmacologically relevant isoforms of carbonic anhydrase were determined by a novel surface plasmon resonance (SPR) biosensor-based approach, using chemodynamic analysis of binding-linked pH-dependent effects. The observed association rates (<i>k</i>_a^obs) were pH-dependent and correlated with the fraction of deprotonated inhibitor and protonated zinc-bound water molecule. The intrinsic association rate constants (<i>k</i>_a^intr) were pH independent and higher than <i>k</i>_a^obs. By contrast, the observed and intrinsic dissociation rate constants were identical and pH-independent, demonstrating that the observed association and dissociation mechanisms are inherently different. A model accounting for the differences between intrinsic and observed rate constants was developed, useful also for other interactions with binding-linked protonation reactions

Kinetically Selective Inhibitors of Human Carbonic Anhydrase Isozymes I, II, VII, IX, XII, and XIII

To get a better understanding of the possibility of developing selective carbonic anhydrase (CA) inhibitors, interactions between 17 benzenesulphonamide ligands and 6 human CAs (full-length CA I, II, VII, and XIII and catalytic domains of CA IX and XII) were characterized using surface plasmon resonance and fluorescent-based thermal shift assays. Kinetics revealed that the strongest binders had subnanomolar affinities with low dissociation rates (i.e., <i>k</i>_d values around 1 × 10^–3 s^–1) or were essentially irreversible. Chemodynamic analysis of the interactions highlighted an intrinsic mechanism of the CA–sulphonamide interaction kinetics and showed that slow dissociation rates were mediated by large hydrophobic contacts. The studied inhibitors demonstrated a high cross-reactivity within the protein family. However, according to chemical phylogenetic analysis developed for kinetic data, several ligands were found to be selective against certain CA isozymes, indicating that it should be possible to develop selective CA inhibitors suitable for clinical use

Identification of a small-molecule ligand of the epigenetic reader protein Spindlin1 via a versatile screening platform

Epigenetic modifications of histone tails play an essential role in the regulation of eukaryotic transcription. Writer and eraser enzymes establish and maintain the epigenetic code by creating or removing posttranslational marks. Specific binding proteins, called readers, recognize the modifications and mediate epigenetic signalling. Here, we present a versatile assay platform for the investigation of the interaction between methyl lysine readers and their ligands. This can be utilized for the screening of small-molecule inhibitors of such protein-protein interactions and the detailed characterization of the inhibition. Our platform is constructed in a modular way consisting of orthogonal in vitro binding assays for ligand screening and verification of initial hits and biophysical, label-free techniques for further kinetic characterization of confirmed ligands. A stability assay for the investigation of target engagement in a cellular context complements the platform. We applied the complete evaluation chain to the Tudor domain containing protein Spindlin1 and established the in vitro test systems for the double Tudor domain of the histone demethylase JMJD2C. We finally conducted an exploratory screen for inhibitors of the interaction between Spindlin1 and H3K4me3 and identified A366 as the first nanomolar small-molecule ligand of a Tudor domain containing methyl lysine reader