Conformational Studies Research Articles

Pushing resolution limits for extracting 1H-1H scalar coupling constants by a resolution-enhanced selective refocusing method.

Nuclear magnetic resonance (NMR) spectroscopy enables one to study molecular structure and dynamics in a noninvasive manner and has long served as a versatile and indispensable analytical tool in physics, chemistry, and biology. Scalar coupling, an essential feature in NMR spectroscopy, provides rich information regarding molecular structure and conformation. The measurement of scalar coupling constants, therefore, constitutes an important issue in NMR spectroscopy. Homonuclear 2D J-resolved spectroscopy is a powerful tool for multiplet analysis and coupling measurement. Recently, a number of phase-sensitive J-resolved methods and selective measuring methods have been developed to facilitate the extraction of coupling constants. However, resolution remains a crucial challenge when extracting small coupling constants or under inhomogeneous fields. In this paper, we present a resolution-enhanced selective refocusing (RESERF) method for the extraction of coupling constants. The effect of magnetic field inhomogeneity can be eliminated, resulting in very narrow linewidths. Therefore, samples with small coupling constants or under inhomogeneous fields can be well analyzed. The RESERF method may be of great value for structural and conformational studies in chemistry and biology.

New Series of Thiazole Derivatives: Synthesis, Structural Elucidation, Antimicrobial Activity, Molecular Modeling and MOE Docking.

Based on the extensive biological activities of thiazole derivatives against different types of diseases, we are interested in the effective part of many natural compounds, so we synthesized a new series of compounds containing di-, tri- and tetrathiazole moieties. The formation of such derivatives proceeded via reaction of 2-bromo-1-(4-methyl-2-(methylamino)thiazol-5-yl)ethan-1-one with heterocyclic amines, o-aminothiophenol and thiosemicarbazone derivatives. The structure and mechanistic pathways for all products were discussed and proved based on spectral results, in addition to conformational studies. Our aim after the synthesis is to investigate their antimicrobial activity against various types of bacteria and fungi species. Preceeding such an investigation, a molecular docking study was carried out with selected conformers, as representative examples, against three pathogen-proteins. This preliminary stage could support the biological application. The potency of these compounds as antimicrobial agents has been evaluated. The results showed that derivatives which have di- and trithiazole rings displayed high activity that exceeds the used standard antibiotic.

Synthesis, conformational analysis and molecular docking studies on three novel dihydropyrimidine derivatives

The dihydropyrimidine (DHPM) derivatives I, II and III were synthesized through Biginelli reaction and characterized by X-ray diffraction and vibrational spectroscopic analyses (Raman and FTIR), whereas a conformational study was conducted. The results show that the DHPM I, II and III belong to the triclinic and monoclinic systems. In a general way, the crystal structure of these compounds are stabilized with N–H⋯O, O–H⋯O and N–H⋯S interactions. The C–H…π and π…π contacts were evaluated using the Hirshfeld surfaces. To evaluate eletronic properties of the DHPM set, theoretical calculations, using B3LYP/6-31G(d,p) method were undertaken. Also, the calculated infrared spectra with some global reactivity descriptors, the molecular electrostatic potential maps and frontier molecular orbital energies were obtained for these compounds. Finally, molecular docking was used to investigate the applicability of DHPM set as a tumor suppressor (L3MBTL1).

Illuminating Biological Interactions with in Vivo Protein Footprinting.

Protein footprinting coupled with mass spectrometry is being increasingly used for the study of protein interactions and conformations. The hydroxyl radical footprinting method, fast photochemical oxidation of proteins (FPOP), utilizes hydroxyl radicals to oxidatively modify solvent accessible amino acids. Here, we describe the further development of FPOP for protein structural analysis in vivo (IV-FPOP) with Caenorhabditis elegans. C. elegans, part of the nematode family, are used as model systems for many human diseases. The ability to perform structural studies in these worms would provide insight into the role of structure in disease pathogenesis. Many parameters were optimized for labeling within the worms including the microfluidic flow system and hydrogen peroxide concentration. IV-FPOP was able to modify several hundred proteins in various organs within the worms. The method successfully probed solvent accessibility similarily to in vitro FPOP, demonstrating its potential for use as a structural technique in a multiorgan system. The coupling of the method with mass spectrometry allows for amino-acid-residue-level structural information, a higher resolution than currently available in vivo methods.

Synthesis, Conformational Analysis and Crystal Structure of New Thioxo, Oxo, Seleno Diastereomeric Cyclophosphamides Containing 1,3,2-dioxaphosphorinane

Thioxo, Oxo and Seleno diastereomeric cyclophosphamides containing 1,3,2- dioxaphosphorinane are prepared by a one-step chemical reaction. Their structural determination is carried out by means of Nuclear Magnetic Resonance NMR (31P, 1 H, 13C) and High-Resolution Mass Spectroscopy (HRMS). The conformational study of diastereomeric products is described. Density Functional Theory (DFT) calculations allowed the identification of preferred conformations. Experimental and calculated 31P, 13C, 1H NMR chemical shifts are compared. The molecular structure of the 2-Benzylamino-5-methyl-5- propyl-2-oxo-1,3,2-dioxaphosphorinane (3d) has been determined by means of crystal Xray diffraction methods.

Structural characterization of organo-montmorillonites prepared from a series of primary alkylamines salts: Mid-IR and near-IR study

Primary alkylammonium cations with progressively increasing chain length from methylammonium (CH3-NH3+) to nonadecylammonium (CH3-(CH2)18-NH3+) were used to prepare unique series of organo-montmorillonites (O-Mts). Infrared spectroscopy in the mid-IR and near-IR regions and XRD were used to characterize samples and to investigate the effect of the chain length on the arrangement of cations confined in montmorillonite (Mt) interlayers. For the first time detailed assignment of the vibration bands in the near-IR region (8000–4000 cm−1) was provided not only for O-Mts but also for selected primary alkylamines and alkylammonium salts, used for their preparation. The IR spectra showed that both, type of head-group, i.e. NH2 vs NH3+, and nature of the anionic counterpart, i.e. chloride vs Mt layer, significantly affected the positions the NH vibration modes as a result of different H-bonds in O-Mts. The alkyl chain length had significant effect on the arrangement of the intercalated cations. With increasing chain length the stretching (νCH2) and first overtone (2νCH2) bands were shifted to lower wavenumbers indicating decreasing amount of disordered gauche in favor of ordered all-trans conformers. Significantly higher shift of the 2νasCH2 band than νasCH2 benefits the NIR region for conformational studies more than MIR.

Synthesis, anti-proliferative activity, theoretical and 1H NMR experimental studies of Morita-Baylis-Hillman adducts from isatin derivatives.

Quaternary or spirocyclic 3-substituted-3-hydroxy-2-oxindole is considered a privileged scaffold. In other words, it is a molecular core present on several compounds with a wide spectrum of biological activities. Among its precursors, activated ketones (isatin nucleus) can be used as interesting starting points to Morita-Baylis-Hillman adducts derivatives, a class of compounds with good cytotoxic potential. In this paper, we present the synthesis, anti-proliferative activity against lung cancer cell line and a theoretical conformational study of 21 of Morita-Baylis-Hillman adducts from isatin derivatives, by DFT quantum chemical calculations, followed by a SAR and QSAR analysis. Besides, an efficient synthetic protocol and good biological activity profile were highlighted interesting observations about 1H NMR experimental spectra, molecular modeling results and crystallographic data available.

Dicyclopenta[ghi,pqr]perylene as a Structural Motif for Bowl-Shaped Hydrocarbons: Synthetic and Conformational Studies.

Dicyclopenta[ghi,pqr]perylene (DCPP) is a substructural fragment on the surface of C70 yet not on C60. Unlike its intensely investigated buckybowl cousins, corannulene and sumanene, DCPP is largely ignored due to the lack of synthetic accessibility. This communication describes the first preparation of a DCPP derivative from bay substituted perylene bis(4-(trifluoromethyl)phenyl)methanol as the key cyclization precursor. Further incorporation of indeno substitutions at peri positions was accomplished through Suzuki-Heck benzannulation. DCPP derivatives 4 adopts a planar structure in crystal. On the contrary, indeno DCPP 5 and bis-indeno DCPP 6 adopt the bowl-shaped conformation in both the solid state and solution. Density functional theory (DFT) calculation reveals that the lowest-energy conformations of 4, 5, and 6 are all bowl-shaped. Nevertheless, small bowl-to-bowl inversion barrier for 4 (3.4 kcal/mol) is overcome by the crystal packing force, which leads to its observed planar structure. However, the bowl-shaped structures of 5 and 6 are affirmed by DFT calculation with intermediate and high bowl-to-bowl inversion barriers (10.4 and 18.5 kcal/mol, respectively).

A New Potent Anti-cancer Corosolic Ester Identified from the Super Miracle Plant Hippophae rhamnoides (Sea buckthorn)

Hippophae rhamnoides L, commonly called Sea buckthorn, is native to Asia and Europe and and known for its nutritional and medicinal values. The aim of the present study was to investigate the anti-cancer constituents of H. rhamnoides. Among the three isolated compounds namely: 1-(2-hydroxynaphthalen-1-yl) ethan-1-one (1), Oleanolic acid (2), and Hippocorosolate (3), compound 3 was a new corosolic ester derivative. The isolated compounds (2 and 3) displayed anticancer activity against lung (NCI-H460) and breast (MCF-7) cancer cell lines with IC50 values of ~3 µM and ~6 µM, respectively. However, compound 1 was active only against breast cancer cells with IC50 value of ~43 µM. These compounds displayed only weak interactions with minor groove of DNA in DNA-ligand conformational studies and therefore, structural DNA damage was not noted in electrophoretic mobility experiments. It was concluded that new compound 2 possessed more potent anticancer activity than that of known compound 3 against lung cancer cell line.

Fluoro-Aryl Substituted α,β2,3-Peptides in the Development of Foldameric Antiparallel β-Sheets: A Conformational Study.

α,β2,3-Disteroisomeric foldamers of general formula Boc(S-Ala-β-2R,3R-Fpg)nOMe or Boc(S-Ala-β-2S,3S-Fpg)nOMe were prepared from both enantiomers of syn H-2-(2-F-Phe)-h-PheGly-OH (named β-Fpg) and S-alanine. Our peptides show two appealing features for biomedical applications: the presence of fluorine, attractive for non-covalent interactions, and aryl groups, crucial for π-stacking. A conformational study was performed, using IR, NMR and computational studies of diastereoisomeric tetra- and hexapeptides containing the β2,3-amino acid in the R,R- and S,S-stereochemistry, respectively. We found that the stability of peptide conformation is dependent on the stereochemistry of the β-amino acid. Combining S-Ala with β-2R,3R-Fpg, a stable extended β-strand conformation was obtained. Furthermore, β-2R,3R-Fpg containing hexapeptide self-assembles to form antiparallel β-sheet structure stabilized by intermolecular H-bonds and π,π-interactions. These features make peptides containing the β2,3-fluoro amino acid very appealing for the development of bioactive proteolytically stable foldameric β-sheets as modulators of protein-protein interaction (PPI).

Conformational and Mechanistic Studies of AlkB and AlkBH2 DNA Demethylases

DNA demethylases play a crucial role in DNA repair mechanisms and have been linked to pathologies. AlkB enzyme and its human homolog AlkBH2 have been very intensively studied both experimentally and computationally. It is well known also that AlkB exhibits broader substrate specificity than AlkBH2. Applying MD and QM/MM methods here we focus on: i) on the differences between the dynamical properties and collective motions between both enzymes: ii) the different impact on the protein environment on the reaction mechanisms in AlkB and AlkBH2. The provided atomistic insight can be used in designing enzyme‐selective inhibitors and demonstrate the power of computational methods to complement experimental studies.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Heterochiral Ala/(αMe)Aze sequential oligopeptides: Synthesis and conformational study.

α-Amino acid residues with a ϕ,ψ constrained conformation are known to significantly bias the peptide backbone 3D structure. An intriguing member of this class of compounds is (αMe)Aze, characterized by an Nα -alkylated four-membered ring and Cα -methylation. We have already reported that (S)-(αMe)Aze, when followed by (S)-Ala in the homochiral dipeptide sequential motif -(S)-(αMe)Aze-(S)-Ala-, tends to generate the unprecedented γ-bend ribbon conformation, as formation of a regular, fully intramolecularly H-bonded γ-helix is precluded, due to the occurrence of a tertiary amide bond every two residues. In this work, we have expanded this study to the preparation and 3D structural analysis of the heterochiral (S)-Ala/(R)-(αMe)Aze sequential peptides from dimer to hexamer. Our conformational results show that members of this series may fold in type-II β-turns or in γ-turns depending on the experimental conditions.

About TFE: Old and New Findings.

The fluorinated alcohol 2,2,2-Trifluoroethanol (TFE) has been implemented for many decades now in conformational studies of proteins and peptides. In peptides, which are often disordered in aqueous solutions, TFE acts as secondary structure stabilizer and primarily induces an α -helical conformation. The exact mechanism through which TFE plays its stabilizing roles is still debated and direct and indirect routes, relying either on straight interaction between TFE and molecules or indirect pathways based on perturbation of solvation sphere, have been proposed. Another still unanswered question is the capacity of TFE to favor in peptides a bioactive or a native-like conformation rather than simply stimulate the raise of secondary structure elements that reflect only the inherent propensity of a specific amino-acid sequence. In protein studies, TFE destroys unique protein tertiary structure and often leads to the formation of non-native secondary structure elements, but, interestingly, gives some hints about early folding intermediates. In this review, we will summarize proposed mechanisms of TFE actions. We will also describe several examples, in which TFE has been successfully used to reveal structural properties of different molecular systems, including antimicrobial and aggregation-prone peptides, as well as globular folded and intrinsically disordered proteins.

Conformational studies of RGDechi peptide by natural-abundance NMR spectroscopy.

Integrins are heterodimeric cell-surface proteins that play important roles during developmental and pathological processes. Diverse human pathologies involve integrin adhesion including thrombotic diseases, inflammation, tumour progression, fibrosis, and infectious diseases. Although in the past decade, novel integrin-inhibitor drugs have been developed for integrin-based medical applications, the structural determinants modulating integrin-ligands recognition mechanisms are still poorly understood, reducing the number of integrin subtype exclusive antagonists. In this scenario, we have very recently showed, by means of chemical and biological assays, that a chimeric peptide (named RGDechi), containing a cyclic RGD motif linked to an echistatin C-terminal fragment, is able to interact with the components of integrin family with variable affinities, the highest for αv β3. Here, in order to understand the mechanistic details driving the molecular recognition mechanism of αv β3 by RGDechi, we have performed a detailed structural and dynamics characterization of the free peptide by natural abundance nuclear magnetic resonance (NMR) spectroscopy. Our data indicate that RGDechi presents in solution an heterogeneous conformational ensemble characterized by a more constrained and rigid pentacyclic ring and a largely unstructured acyclic region. Moreover, we propose that the molecular recognition of αv β3 integrin by RGDechi occurs by a combination of conformational selection and induced fit mechanisms. Finally, our study indicates that a detailed NMR characterization, by means of natural abundance 15 N and 13 C, of a mostly unstructured bioactive peptide may provide the molecular basis to get essential structural insights into the binding mechanism to the biological partner.

Bicyclic Pyrrolidine-Isoxazoline γ Amino Acid: A Constrained Scaffold for Stabilizing α-Turn Conformation in Isolated Peptides.

Unnatural amino acids have tremendously expanded the folding possibilities of peptides and peptide mimics. While α,α-disubstituted and β-amino acids are widely studied, γ-derivatives have been less exploited. Here we report the conformational study on the bicyclic unnatural γ amino acid, 4,5,6,6a-tetrahydro-3aH-pyrrolo[3,4-d]isoxazole-3-carboxylic acid 1. In model peptides, the (+)-(3aR6aS)-enantiomer is able to stabilize α-turn conformation when associated to glycine, as showed by 1H-NMR, FT-IR, and circular dichroism experiments, and molecular modeling studies. α-turn is a structural motif occurring in many biologically active protein sites, although its stabilization on isolated peptides is quite uncommon. Our results make the unnatural γ-amino acid 1 of particular interest for the development of bioactive peptidomimetics.

Extended Diethylglycine Homopeptides Formed by Desulfurization of Their Tetrahydrothiopyran Analogues.

Diethylglycine (Deg) homopeptides adopt the rare 2.05-helical conformation, the longest three-dimensional structure that a peptide of a given sequence can adopt. Despite this unique conformational feature, Deg is rarely used in peptide design because of its poor reactivity. In this paper, we show that reductive desulfurization of oligomers formed from more reactive tetrahydrothiopyran-containing precursors provides a practical way to build the longest Deg homopeptides so far made, and we detail some conformational studies of the Deg oligomers and their heterocyclic precursors.

Hybrid Supramolecules for Azolium-Linked Cyclophane Immobilization and Conformation Study: Synthesis, Characterization, and Photocatalytic Degradation.

In this paper, one imidazole macrocyclic divalent cation with a flexible configuration was chosen with the aim of the immobilization of its conformation. Six novel organic–inorganic hybrid supramolecules {([syn/anti-did](CdI4)(C2H3N)} (1), {([syn/anti-did](HgI4)(C2H3N)} (2), {([syn/syn-did](Pb2I8)(PbI5)} (3), {([syn/syn-did](Bi2I8)(BiI5)} (4), {([syn/syn-did](Ag4Br6)}n (5), and {([syn/anti-did](Ag2I4)} (6) [did2+ = (12z, 52z)-11H, 51H-1, 5(1, 3)-diimidazol-3-iuma-3, 7(1, 2)-dibenzenacyclooctaphane-13, 53-diium] have been synthesized through the self-assembly reaction of did2+ with different metals under solvothermal reaction. These compounds have been unambiguously confirmed by powder X-ray diffraction, IR, thermogravimetric and X-ray single-crystal diffraction. Crystallographic analysis shows that the anions of compounds 1 and 2 are isostructural with mononuclear anion structure, the anions of compounds 3, 4, and 6 are of binuclear structure, and compound 5 anion is a one-dimensional anionic chain structure. Imidazole cyclophanes of organic cations may exist in a syn/syn-conformation in 3–5 or syn/anti conformation in 1, 2, 6, but tilting angles in compounds 1–6 were different. This article described the photocatalytic degradation of organic dye in waste water and the optical band gap of compounds 1–6. The calculated semiconductor band gap of compounds 1–6 is smaller than that of TiO2, so compounds 1–6 have semiconductor properties. Compound 1 has the smallest band gap value, and the photocatalytic performance is the best.

Synthesis, thermal stability, vibrational spectra and conformational studies of novel dicationic meta-xylyl linked bis-1-methylimidazolium ionic liquids

In recent years, dicationic ionic liquids have been subject of a strongly growing research effort for their interesting and important properties that suggest them to be potential candidate for many applications. Therefore, the knowledge about the synthesis, thermal stability, vibrational spectra and conformational behavior of these compounds is an important subject of study. The objective of the work presented in this paper is to investigate the influence of the substituent relative meta-position of dicationic ionic liquids (ILs), namely meta-xylyl linked bis-1-methylimidazolium ILs, on thermal stability, vibrational spectra and conformational studies. In the first step, four new meta-xylyl linked bis-imidazolium [m-C6H4(CH2ImMe)2+] ionic liquid (ILs) containing the dichloride [Cl−]2, di(bis(trifluoromethylsulfonyl) Imide) [(CF3SO2)2N−]2, bis(tetrafluoroborate) [BF4−]2 and bis (hexafluorophosphate) [PF6−]2 anions were synthesized. The obtained dicationic ILs were characterized by 1H NMR, 13C NMR, 19F NMR, 31P NMR and FT-IR spectroscopy. Also, their thermal properties were determined and compared. The thermal behavior confirmed that [m-C6H4(CH2ImMe)2+] dicationic ILs containing [(CF3SO2)2N−]2 and [PF6−]2 are more stable that the dichloride [Cl−]2, and bis(tetrafluoroborate) [BF4−]2 dicationic ILs, and they show a good thermal stability up to 340 °C which makes them suitable for thermal application. In the next step, a computational study of the conformers of [m-C6H4(CH2ImMe)2], by means of DFT calculations with the 6-31G** basis set and the B3LYP theory was performed. Additionally, vibrational spectroscopy studies were conducted by infrared (FTIR) and Raman (FT-Raman) spectroscopy on the four dicationic ILs, differing for their anions. Finally a computational investigation of the interactions between anions and cation is presented.

Study of molecular structure and hydrogen bond interactions in dipfluzine-benzoic acid (DIP-BEN) cocrystal using spectroscopic and quantum chemical method.

The purpose of this article is to predict the molecular structure of the cocrystal of dipfluzine-benzoic acid (DIP-BEN) through computational approach (DFT calculations) and validate it using vibrational spectroscopic studies. The molecular structure of the DIP-BEN cocrystal has been predicted by forming models on the basis of the active sites available to form H-bonds between dipfluzine (DIP) and benzoic acid (BEN). Conformational study has been performed and potential energy surface scans are plotted around the flexible bonds of the cocrystal molecule and three stable conformers have been obtained. Quantum theory of atoms in molecules (QTAIM) explains that all the interactions are medium and partially covalent in nature. Natural bond orbital analysis of the second order perturbation theory of the Fock matrix suggests that interactions LP (2) O2 → σ*(O74H75) and LP (2) F1 → σ* (O89H90) are responsible for the stabilization of the molecule. The HOMO and LUMO energies and electronic charge transfer (ECT) confirms that charge flows from BEN to DIP. Global reactivity descriptor parameters suggest that DIP-BEN cocrystal is softer, thus more reactive in comparison to DIP. Local reactivity descriptor parameter is used to predict reactive sites of the cocrystal. The experimental and theoretical results support the formation of cocrystal through strong hydrogen bond (O89H90⋯F1 and O74H75⋯O2) interactions present in cocrystal.

Conformational dynamics study on human γS-crystallin as an efficient route to childhood blindness

Single point mutants of human γS-crystallin cause dominant congenital cataracts, a recent one of which involves the substitution of highly conserved glycine at 57th position with a bulkier tryptophan. Our high-resolution 3D structure of this G57W mutant (abbreviated hereafter as γS-G57W), reported recently revealed site-specific structural perturbations with higher aggregation and lower stability compared to its wild-type; a structural feature associated with important functional and therapeutic consequences. In this communication, we report for the first time, residue resolved conformational dynamics in both γS-WT and γS-G57W using solution NMR spectroscopy, and suggest how these differences could crucially affect the biochemistry of the mutant. Guided by our critical structural investigations, extensive conformational dynamics and biophysical studies presented here show that loss of structural stability arises from enhanced dynamics in Greek key motif 2 inducing flexibility in the N-terminal domain as opposed to its structurally unperturbed C-terminal counterpart. NMR spectral density correlations and internal dynamics comparisons with the wild-type suggest that the overall thermodynamic instability propagates from the mutated N-terminal β4-β5 loop providing a residue level understanding of the structural changes associated with this early onset of lens opacification. Our results highlight the vital role of conserved Greek key motifs in conferring structural stability to crystallins and provide crucial molecular insights into crystallin aggregation in the eye lens, which triggers cataract formation in children. Overall, this critical study provides a residue level understanding of how conformational changes affect the structure and function of crystallins in particular and proteins in general, during health and disease.