The Role of Rare-Earth Atoms in the Anisotropy and Antiferromagnetic Exchange Coupling at a Hybrid Metal-Organic Interface.

Magnetic anisotropy and magnetic exchange interactions are crucial parameters that characterize the hybrid metal-organic interface, a key component of an organic spintronic device. It is shown that the incorporation of 4f RE atoms to hybrid metal-organic interfaces of CuPc/REAu2 type (RE = Gd, Ho) constitutes a feasible approach toward on-demand magnetic properties and functionalities. The GdAu2 and HoAu2 substrates differ in their magnetic anisotropy behavior. Remarkably, the HoAu2 surface promotes the inherent out-of-plane anisotropy of CuPc, owing to the match between the anisotropy axis of substrate and molecule. Furthermore, the presence of RE atoms leads to a spontaneous antiferromagnetic exchange coupling at the interface, induced by the 3d-4f superexchange interaction between the unpaired 3d electron of CuPc and the 4f electrons of the RE atoms. It is shown that 4f RE atoms with unquenched quantum orbital momentum ( L $L$ ), as it is the case of Ho, induce an anisotropic interfacial exchange coupling.

Thermodynamic and structural investigation of the interaction of quaternized 2,3-octakis-[(2-mercaptopyridine)phthalocyaninato] copper (II) sulfate (CuPc) with parallel and hybrid type G-quadruplex.

G-quadruplexes are important drug targets and get attention due to their existence in telomere, ribosomal DNA, promoter regions of some oncogenes, and the untranslated regions of mRNA. Due to the biological roles of G-quadruplexes, investigating of the G-quadruplex-small molecule interaction is essential. The primary motivation for these studies is the possibility of inhibiting cell functions associated with G-quadruplex sequences by binding with small molecules. Targeting the small molecules to desired tissue with the G-quadruplex vehicles is the second important goal of the G-quadruplex-small molecule interaction studies. In the present study, the new peripherally 2-mercaptopyridine octasubstituted copper(II) phthalocyanine and its quaternized derivative (CuPc) were synthesized and characterized by elemental analysis FT-IR, UV-Vis, and mass spectra. The excellent solubility of CuPc in water is essential for its transport in the organism. Because of this feature, its affinity toward G-quadruplex forming aptamers, AS1411, Tel21, and Tel45, was investigated. The UV-Vis spectrophotometric titration data confirmed the prevention of aggregation upon interaction with G-quadruplex, which is very important for biomedical applications. The CD spectroscopic analyses and binding stoichiometry confirmed the "end stacking" model for interaction of AS1411 with CuPc. The interaction of CuPc caused the equilibrium shift from hybrid conformation to antiparallel conformation for Tel21 and Tel45. The isothermal titration calorimeter (ITC) was used for the determination of thermodynamic parameters. The thermodynamic data of the interaction was fitted well with the one-site model. The negative values of Gibbs free energy change confirmed the spontaneous nature of the reactions. Besides, the negative values of enthalpy change and entropy change proved that the nature of processes was "enthalpy driven." The interaction stoichiometry was 2 for AS1411 and Tel21 and 1.5 for Tel45. The binding constants were 1.3(±0.3) × 105 , 3.2(±0.4) × 105 , and 1.1(±0.3) × 105 M-1 , which were at the level of ethidium bromide intercalation binding constant given in the literature. The DNA polymerase stop assay further supported the interaction of CuPc with G-quadruplex DNA. The experimental results confirm that the CuPc has a potential photosensitizer behaviour for photodynamic therapy.

Biological Evaluation and Binding Mechanism of 5-HT7 Specific Arylpiperazinyl-Alkyl Benzothiazolone: Radiobiology and Photo-physical Studies.

Diversely substituted methoxy derivatives of arylpiperazinyl-alkyl benzothiazolone has been evaluated as specific probe for 5HT7. To determine the best methoxy derivative for 5HT7 receptor affinity, we synthesised a number of 2-benzothiazolone arylalkyl piperazine derivatives. In-vitro/vivo studies with C-2 substituted [11C]ABT showed 5HT7 specific binding. The radiochemical purity of [11C]ABT was found to be more than 99% with radiochemical stability persistence for more than 1.5 hr at 25 °C. The interaction of BSA and ABT has been analysed by photophysical studies for better understanding of properties such as adsortion, distribution, metabolism and elemination (ADME). The interaction between ABT and BSA was analyzed by using the UV-vis and fluorescence spectra. UV-vis spectra analyzed the changes in primary structure of BSA on its interaction with ABT. ABT showed quenched fluorescence emission intensity of tryptophan residues in BSA via static quenching mechanism. This study might help to understand how ABT binds to serum protein or subsequently to know the ADME of this drug candidate.

Optical Relay Sensing of Cryptochiral Alcohols Displaying α-, ß-, γ- and δ-Stereocenters or Chirality by Virtue of Isotopic Substitution.

A reaction-based optical relay sensing strategy that enables accurate determination of the concentration and enantiomeric ratio (er) of challenging chiral alcohols exhibiting stereocenters at the α-, ß-, γ- or even δ-position or hard-to-detect cryptochirality arising from H/D substitution is described. This unmatched application scope is achieved with a conceptually new sensing approach by which the alcohol moiety is replaced with an optimized achiral sulfonamide chromophore to minimize the distance between the covalently attached chiroptical reporter unit and the stereogenic center in the substrate. The result is a remarkably strong, red-shifted CD induction that increases linearly with the sample er. The CD sensing part of the tandem assay is seamlessly coupled to a redox reaction with a quinone molecule to generate a characteristic UV response that is independent of the enantiopurity of the alcohol and thus allows determination of the total analyte concentration. The robustness and utility of the CD/UV relay are further verified by chromatography-free asymmetric reaction analysis with small aliquots of crude product mixtures, paving the way toward high-throughput chiral compound screening workflows which is a highly sought-after goal in the pharmaceutical industry.

Conformational change in an engineered biliverdin-binding cyanobacteriochrome during the photoconversion process.

Cyanobacteriochromes (CBCRs) derived from cyanobacteria are linear-tetrapyrrole-binding photoreceptors related to the canonical red/far-red reversible phytochrome photoreceptors. CBCRs contain chromophore-binding cGMP-specific phosphodiesterase/adenylate cyclase/FhlA (GAF) domains that are highly diverse in their primary sequences and are categorized into many subfamilies. Among this repertoire, the biliverdin (BV)-binding CBCR GAF domains receive considerable attention for their in vivo optogenetic and bioimaging applications because BV is a mammalian intrinsic chromophore and can absorb far-red light that penetrates deep into the mammalian body. The typical BV-binding CBCR GAF domain exhibits reversible photoconversion between far-red-absorbing dark-adapted and orange-absorbing photoproduct states. Herein, we applied various biochemical and spectral studies to identify the details of the conformational change during this photoconversion process. No oligomeric state change was observed, whereas the surface charge would change with a modification of the α-helix structures during the photoconversion process. Combinatorial analysis using partial protease digestion and mass spectrometry identified the region where the conformational change occurred. These results provide clues for the future development of optogenetic tools.

Effect of Neurodegenerative Mutations in the NEFL Gene on Thermal Denaturation of the Neurofilament Light Chain Protein.

Effects of E90K, N98S, and A149V mutations in the light chain of neurofilaments (NFL) on the structure and thermal denaturation of the NFL molecule were investigated. By using circular dichroism spectroscopy, it was shown that these mutations did not lead to the changes in α-helical structure of NFL, but they caused noticeable effects on the stability of the molecule. We also identified calorimetric domains in the NFL structure by using differential scanning calorimetry. It was shown that the E90K replacement leads to the disappearance of the low-temperature thermal transition (domain 1). The mutations cause changes in the enthalpy of NFL domains melting, as well as lead to the significant changes in the melting temperatures (Tm) of some calorimetric domains. Thus, despite the fact that all these mutations are associated with the development of Charcot-Marie-Tooth neuropathy, and two of them are even located very close to each other in the coil 1A, they affect differently structure and stability of the NFL molecule.

Strain Tunability of Perpendicular Magnetic Anisotropy in van der Waals Ferromagnets VI3.

Layered ferromagnets with strong magnetic anisotropy energy (MAE) have special applications in nanoscale memory elements in electronic circuits. Here, we report a strain tunability of perpendicular magnetic anisotropy in van der Waals (vdW) ferromagnets VI3 using magnetic circular dichroism measurements. For an unstrained flake, the M-H curve shows a rectangular-shaped hysteresis loop with a large coercivity (1.775 T at 10 K) and remanent magnetization. Furthermore, the coercivity can be enhanced to a maximum of 2.6 T under a 3.8% external in-plane tensile strain. Our DFT calculations show that the increased MAE under strain contributes to the enhancement of coercivity. Meanwhile, the strain tunability on the coercivity of CrI3, with a similar crystal structure, is limited. The main reason is the strong spin-orbit coupling in V3+ in VI6 octahedra in comparison with that in Cr3+. The strain tunability of coercivity in VI3 flakes highlights its potential for integration into vdW heterostructures.

Advantages of Induced Circular Dichroism Spectroscopy for Qualitative and Quantitative Analysis of Solution-Phase Cyclodextrin Host-Guest Complexes.

The presence of a chiral or chirally perturbed chromophore in the molecule under investigation is a fundamental requirement for the appearance of a circular dichroism (CD) spectrum. For native and for most of the substituted cyclodextrins, this condition is not applicable, because although chiral, cyclodextrins lack a chromophore group and therefore have no characteristic CD spectra over 220 nm. The reason this method can be used is that if the guest molecule has a chromophore group and this is in the right proximity to the cyclodextrin, it becomes chirally perturbed. As a result, the complex will now provide a CD signal, and this phenomenon is called induced circular dichroism (ICD). The appearance of the ICD spectrum is clear evidence of the formation of the complex, and the spectral sign and intensity is a good predictor of the structure of the complex. By varying the concentration of cyclodextrin, the ICD signal changes, resulting in a saturation curve, and from these data, the stability constant can be calculated for a 1:1 complex. This article compares ICD and NMR spectroscopic and molecular modeling results of cyclodextrin complexes of four model compounds: nimesulide, fenbufen, fenoprofen, and bifonazole. The results obtained by the different methods show good agreement, and the structures estimated from the ICD spectra are supported by NMR data and molecular modeling.

Transformable Plasmonic Helix with Swinging Gold Nanoparticles.

Control over multiple optical elements that can be dynamically rearranged to yield substantial three-dimensional structural transformations is of great importance to realize reconfigurable plasmonic nanoarchitectures with sensitive and distinct optical feedback. In this work, we demonstrate a transformable plasmonic helix system, in which multiple gold nanoparticles (AuNPs) can be directly transported by DNA swingarms to target positions without undergoing consecutive stepwise movements. The swingarms allow for programmable AuNP translocations in large leaps within plasmonic nanoarchitectures, giving rise to tailored circular dichroism spectra. Our work provides an instructive bottom-up solution to building complex dynamic plasmonic systems, which can exhibit prominent optical responses through cooperative rearrangements of the constituent optical elements with high fidelity and programmability.

Biophysical characterization of full-length oleosin in dodecylphosphocholine micelles.

Oleosin is a hydrophobic protein that punctuates the surface of plant seed lipid droplets, which are 20 nm-100 µm entities that serve as reservoirs for high-energy metabolites. Oleosin is purported to stabilize lipid droplets, but its exact mechanism of stabilization has not been established. Probing the structure of oleosin directly in lipid droplets is challenging due to the size of lipid droplets and their high degree of light scattering. Therefore, a medium in which the native structure of oleosin is retained, but is also amenable to spectroscopic studies is needed. Here, we show, using a suite of biophysical techniques, that dodecylphosphocholine micelles appear to support the tertiary structure of the oleosin protein (i.e., hairpin conformation) and render the protein in an oligomeric state that is amenable to more sophisticated biophysical techniques such as NMR.

Structural analysis of the peptides temporin-Ra and temporin-Rb and interactions with model membranes.

The skin of amphibians is widely exploited as rich sources of membrane active peptides that differ in chain size, polypeptide net charge, secondary structure, target selectivity and toxicity. In this study, two small antimicrobial peptides, temporin-Ra and temporin-Rb, originally isolated from the skin of the European marsh frog (Rana ridibunda), described as active against pathogen bacteria and presenting low toxicity to eukaryotic cells were synthesized and had their physicochemical properties and mechanism of action investigated. The temporin peptides were examined in aqueous solution and in the presence of membrane models (lipid monolayers, micelles, lipid bilayers and vesicles). A combined approach of bioinformatics analyses, biological activity assays, surface pressure measurements, synchrotron radiation circular dichroism spectroscopy, and oriented circular dichroism spectroscopy were employed. Both peptides were able to adsorb at a lipid-air interface with a negative surface charge density, and efficiently disturb the lipid surface packing. A disorder-to-helix transition was observed on the secondary structure of both peptides when either in a non-polar environment or interacting with model membranes containing a negative net charge density. The binding of both temporin-Ra and temporin-Rb to membrane models is modulated by the presence of negatively charged lipids in the membrane. The amphipathic helix induced in temporin-Ra is oriented parallel to the membrane surface in negatively charged or in zwitterionic lipid bilayers, with no tendency for realignment after binding. Temporin-Rb, instead, assumes a ß-sheet conformation when deposited into oriented stacked lipid bilayers. Due to their short size and simple composition, both peptides are quite attractive for the development of new classes of peptide-based anti-infective drugs.

Influence of the heme distal pocket on nitrite binding orientation and reactivity in Sperm Whale myoglobin.

Nitrite binding to recombinant wild-type Sperm Whale myoglobin (SWMb) was studied using a combination of spectroscopic methods including room-temperature magnetic circular dichroism. These revealed that the reactive species is free nitrous acid and the product of the reaction contains a nitrite ion bound to the ferric heme iron in the nitrito- (O-bound) orientation. This exists in a thermal equilibrium with a low-spin ground state and a high-spin excited state and is spectroscopically distinct from the purely low-spin nitro- (N-bound) species observed in the H64V SWMb variant. Substitution of the proximal heme ligand, histidine-93, with lysine yields a novel form of myoglobin (H93K) with enhanced reactivity towards nitrite. The nitrito-mode of binding to the ferric heme iron is retained in the H93K variant again as a thermal equilibrium of spin-states. This proximal substitution influences the heme distal pocket causing the pKa of the alkaline transition to be lowered relative to wild-type SWMb. This change in the environment of the distal pocket coupled with nitrito-binding is the most likely explanation for the 8-fold increase in the rate of nitrite reduction by H93K relative to WT SWMb.

Crystal Structure of H227A Mutant of Arginine Kinase in Daphnia magna Suggests the Importance of Its Stability.

Arginine kinase (AK) plays a crucial role in the survival of Daphnia magna, a water flea and a common planktonic invertebrate sensitive to water pollution, owing to the production of bioenergy. AK from D. magna (DmAK) has four highly conserved histidine residues, namely, H90, H227, H284, and H315 in the amino acid sequence. In contrast to DmAK WT (wild type), the enzyme activity of the H227A mutant decreases by 18%. To identify the structure-function relationship of this H227A mutant enzyme, the crystal 3D X-ray structure has been determined and an unfolding assay using anilino-1-naphthalenesulfonic acid (ANS) fluorescence has been undertaken. The results revealed that when compared to the DmAK WT, the hydrogen bonding between H227 and A135 was broken in the H227A crystal structure. This suggests that H227 residue, closed to the arginine binding site, plays an important role in maintaining the structural stability and maximizing the enzyme activity through hydrogen bonding with the backbone oxygen of A135.

Spectroscopic Studies of Mononuclear Molybdenum Enzyme Centers.

A concise review is provided of the contributions that various spectroscopic methods have made to our understanding of the physical and electronic structures of mononuclear molybdenum enzymes. Contributions to our understanding of the structure and function of each of the major families of these enzymes is considered, providing a perspective on how spectroscopy has impacted the field.

Red-Beet Betalain Pigments Inhibit Amyloid-ß Aggregation and Toxicity in Amyloid-ß Expressing Caenorhabditis elegans.

Betalain pigments are mainly produced by plants belonging to the order of Caryophyllales. Betalains exhibit strong antioxidant activity and responds to environmental stimuli and stress in plants. Recent reports of antioxidant, anti-inflammatory and anti-cancer properties of betalain pigments have piqued interest in understanding their biological functions. We investigated the effects of betalain pigments (betanin and isobetanin) derived from red-beet on amyloid-ß (Aß) aggregation, which causes Alzheimer's disease. Non-specific inhibition of Aß aggregation against Aß40 and Aß42 by red-beet betalain pigments, in vitro was demonstrated using the thioflavin t fluorescence assay, circular dichroism spectroscopy analysis, transmission electron microscopy and nuclear magnetic resonance (NMR) analysis. Furthermore, we examined the ability of red-beet betalain pigments to interfere with Aß toxicity by using the transgenic Caenorhabditis elegans model, which expresses the human Aß42 protein intracellularly within the body wall muscle. It responds to Aß-toxicity with paralysis and treatment with 50 µM red-beet betalain pigments significantly delayed the paralysis of C. elegans. These results suggest that betalain pigments reduce Aß-induced toxicity.

Rapid preparation of nanodiscs for biophysical studies.

Nanodiscs, which are disc-shaped entities that contain a central lipid bilayer encased by an annulus of amphipathic helices, have emerged as a leading native-like membrane mimic. The current approach for the formation of nanodiscs involves the creation of a mixed-micellar solution containing membrane scaffold protein, lipid, and detergent followed by a time consuming process (3-12 h) of dialysis and/or incubation with sorptive beads to remove the detergent molecules from the sample. In contrast, the methodology described herein provides a facile and rapid procedure for the preparation of nanodiscs in a matter of minutes (<15 min) using Sephadex® G-25 resin to remove the detergent from the sample. A panoply of biophysical techniques including analytical ultracentrifugation, dynamic light scattering, gel filtration chromatography, circular dichroism spectroscopy, and cryogenic electron microscopy were employed to unequivocally confirm that aggregates formed by this method are indeed nanodiscs. We believe that this method will be attractive for time-sensitive and high-throughput experiments.

Effect of setting data collection parameters on the reliability of a circular dichroism spectrum.

Circular dichroism (CD) spectroscopy is a well-established biophysical technique used to investigate the structure of molecules. The analysis of a protein CD spectrum depends on the quality of the original CD data, which can be affected by the sample purity, background absorption of the additives/solvent/buffer, the choice of the parameters used for data collection, etc. In this paper, the CD spectrum of myoglobin was used as a model to exploit how variations on each data collection parameter could affect the final protein CD spectrum and, the subsequent effect of them on the quantitative analysis of protein secondary structure. Bioinformatics analysis carried out with SESCA package and PDBMD2CD server predicted a theoretical myoglobin CD spectrum, and a Monte Carlo-like model was implemented to estimate the uncertainty in secondary structure predictions performed with CDSSTR, Selcon 3 and ContinLL algorithms. An inappropriate choice of data collection parameters can lead to a misinterpretation of the CD data in terms of the protein structural content.

Novel anoplin-based (lipo)-peptide models show potent antimicrobial activity.

The subject of this study is the synthesis and biological evaluation of anoplin-based (Gly-Leu-Leu3 -Lys-Arg5 -Ile-Lys-Thr8 -Leu-Leu-NH2 )-designed (lipo)-peptides, aiming at the development of new antibiotic substances. The design of synthetic compounds based on natural bioactive molecules is an optimistic strategy for the development of new pharmaceutics. Antimicrobial peptides (AMPs) and (lipo)-peptides are two classes of promising compounds, with characteristics that allow them to express their activity by differentiated mechanisms of action. On this basis, anoplin, a natural AMP, was used as a scaffold to design five peptides and seven lipopeptide analogs of them. Substitutions were made on residues Leu3 and Arg5 of the interphase and on Thr8 of the polar phase, as well as N-terminus conjunctions with octanoic and decanoic acid. The outcome of the biological evaluation revealed that some analogs might have substantial clinical potential. Specifically, Ano 1-F, Ano 3-F, Ano 4-C10 , and Ano 5-F are strongly active against Gram-negative bacteria at minimum inhibitory concentration (MIC) values of 3 µg/ml, while Ano 4-F is active against Gram-positive bacteria at 1 µg/ml. Ano 2-C10 , C10 -Gly-Leu-Lys3 -Lys-Ile5 -Ile-Lys-Lys8 -Leu-Leu-NH2 , is the most promising compound (MIC = 0.5 µg/ml) for the development of new pharmaceutics. The conformational features of the synthetic peptides were investigated by circular dichroism spectroscopy, and their physicochemical parameters were calculated. Our study shows that appropriate substitutions in the anoplin sequence in combination with Nα -acylation may lead to new effective AMPs.

Examination of the Antimicrobial Peptide Myticalin A6 Active Site.

In 2017, Leoni et al. reported myticalins as novel cationic linear antimicrobial peptides obtained from marine mussels. The authors focused on myticalin A6 (29 amino acids), which has a relatively short chain length among myticalins and contains a repeating X-proline(Pro)-arginine (Arg) sequence in its structure. We investigated the antimicrobial activity of myticalin A6 against Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus (S. aureus). Fragment derivatives of myticalin A6 were synthesized, and the site required for expression of antimicrobial activity was examined. To investigate the structure-antimicrobial activity relationship of myticalin A6, short-chain derivatives and partially substituted derivatives were synthesized, and the antimicrobial activity was measured. Furthermore, some cyclized derivatives were synthesized and examined for antimicrobial activity. Circular dichroism (CD) spectroscopy of myticalin A6 and its derivatives was carried out to evaluate the secondary structure. Myticalin A6 exhibited an antimicrobial activity of 1.9 µM against S. aureus. Myticalin A6 (3-23)-OH (21 amino acids) exhibited an antimicrobial activity of 2.4 µM against S. aureus, suggesting that the X-Pro-Arg repeat sequence is important for antimicrobial activity. Derivatives with different CD measurement results from myticalin A6 (3-23)-OH exhibited decreased activity. The myticalin A6 (3-23)-OH derivative in which all Arg residues were replaced with lysine (Lys) residues exhibited reduced antimicrobial activity against S. aureus. We succeeded in synthesizing cyclic derivatives using 9-fluorenylmethoxycarbonyl (Fmoc)-aspartic acid (Asp)(Wang resin)-[2-phenylisopropyl ester (OPis)], but the yield of derivatives with 21 amino acids was decreased. The myticalin derivatives synthesized in this study did not exhibit any enhancement in antimicrobial activity due to cyclization.

Vibrational circular dichroism of d-amino acid-containing peptide NdWFamide in the crystal form.

Microcrystals of l-Asn-d-Trp-l-Phe-NH2 (NdWFamide), a tripeptide derived from Aplysia kurodai that exhibits invertebrate cardiac activity, were evaluated by vibrational circular dichroism (VCD). The chirality of the tryptophan residue at the second position in NdWFamide was associated with the conformation and biological characteristics. The VCD spectrum of NdWFamide was a mirror image of its enantiomer; however, it was significantly different from that of its diastereomer, NWFamide, which is its precursor. The obtained VCD signals of NdWFamide were in good agreement with the VCD signals that were calculated based on the optimized aggregates of NdWFamide, which formed a helical-like backbone conformation. The evaluation of the VCD results revealed the conformation of NdWFamide in the crystalline state and succeeded in distinguishing its stereoisomers. Therefore, this study demonstrates VCD as a useful method for the structural analysis of naturally occurring d-amino acid-containing peptides.