An efficient use of the WATERGATE W5 sequence for observing a ligand binding with a protein receptor

An efficient pulse sequence for observing a ligand binding with a receptor has been developed by incorporating the WATERGATE W5 sequence. In the conventional water ligand observed via gradient spectroscopy (WaterLOGSY) techniques, the water resonance is selectively excited using, e.g. the double-pulsed field gradient spin-echo (DPFGSE) sequence at the initial portion of pulse sequence. In the current version, the modified WATERGATE W5 sequence is incorporated at the initial portion of the pulse sequence, and the resonance at the water frequency can be selectively reserved by the modified WATERGATE W5 sequence. The efficiency of ligand-observed NMR screening techniques has been demonstrated using the human serum albumin (HSA)-tryptophan complex.     

Aroma WaterLOGSY: a fast and sensitive screening tool for drug discovery

One-dimensional NMR spectroscopy has proven to be a powerful technique for screening compound libraries in drug discovery. We report a novel water ligand-observed gradient spectroscopy (WaterLOGSY) pulse sequence, named Aroma WaterLOGSY, that selectively detects aromatic WaterLOGSY signals from compounds or ligands. In the Aroma WaterLOGSY, water magnetization is untouched after water excitation and utilizes the whole period of the remaining pulse sequence to relax back to the +z direction. Due to the phase cycling design, the water magnetization is allowed to relax for the period of two full scans before it gets inverted again. Therefore, the recycle delay can be significantly shortened. Within similar experimental time, Aroma WaterLOGSY shows approximately two times higher sensitivity than the standard scheme. This method also allows the use of non-deuterated reagents, thereby accelerating experimental set-up time for ligand-binding studies.     

Application of NMR SHAPES screening to an RNA target

Several NMR screening techniques have been developed in recent years to aid in the identification of lead drug compounds. These NMR methods have traditionally been used for protein targets, and here we examine their applicability for an RNA target. We used the SHAPES compound library to test three different NMR screening methodologies: the saturation transfer difference (STD), the 2D trNOESY, and the WaterLOGSY experiments. We found that the WaterLOGSY experiment was the most sensitive method for our RNA target, the P4P6 domain of the Tetrahymena thermophila Group I intron. Using the WaterLOGSY experiment, we found that 23 of the 112 SHAPES compounds interact with P4P6. To identify which of these 23 hits bind through nonspecific interactions, we counterscreened with a linear duplex RNA control and identified one of the SHAPES compounds as interacting with P4P6 specifically. We thus demonstrated that the WaterLOGSY experiment in combination with the SHAPES compound library can be used to efficiently find RNA binding lead compounds.     

环三聚磷腈多齿配体的合成及DNA的切割活性

为了得到具有核酸切割功能的人工核酸酶, 设计合成了5种环三聚磷腈多齿配体, 并初步检测了其对DNA的切割活性. 目标化合物的结构由IR, 1H NMR, 31P NMR, 13C NMR和ESI-MS确认. 在生理条件下对pUC19 DNA切割活性的初步实验结果表明, 在化合物5a～5e的Cu(Ⅱ)配合物存在下, 保温24 h后, pUC19 DNA由Form Ⅰ断裂为Form Ⅱ, 即合成目标化合物有明显的DNA切割活性. 同时, 考察了配合物5b+Cu在不同时间下对DNA的切割活性的影响.     

Probing the dimensions of semi-rigid inner functionalised U-shaped bis-porphyrin cavities

The synthesis of a series of open U-shaped bis-porphyrin cavity molecules is described, with bridged bicyclic backbones to confer rigidity, and a pendant substituted aromatic probe unit suspended on the inside of the cavity. The dimensions and flexibility of the bis-zincporphyrin cavity were probed using several different techniques. Initially the molecular ruler concept was employed, using flexible bidentate ligands as guests with a range of possible linear dimensions. Secondly, NMR methods were employed with rigid bidentate ligand guests of fixed lengths, and thirdly diffusion based NMR methods were utilised. The range of inter-porphyrin distances estimated by these methods suggests that these types of open-cavity systems have a surprising degree of flexibility.     

Cysteine and Methionine Derivatives of Aminophenol Ligands: Synthesis,Iron Complexation,Magnetic, Electronic and Redox Investigation

Two new Cysteine and Methionine derivatives of aminophenol ligands (HL^Cys and HL^Met) were synthesized by a convenient procedure. The ligands were characterized by ¹H NMR, ¹³C NMR and IR spectroscopies, ESI‐MS, elemental analysis and optical activity measurements. Iron(III) complexes (FeL^Cys and FeL^Met) of these ligands were synthesized and characterized by spectroscopic, magnetic susceptibility studies and cyclic voltammetry techniques. The molecular structure of FeL^Cys and FeL^Met determined by ESI‐MS consist of two ligands coordinated to Fe(III) centers. The magnetic susceptibility measurement indicates the monomeric complexes with paramagnetic properties. Both complexes undergo metal‐centred reduction, and ligand‐centred oxidation.     

Synthesis and properties of metal-ligand complexes with endohedral amine functionality

A series of tetracationic M(2)L(4) palladium-pyridyl complexes with endohedral amine functionality have been synthesized. The complexes were analyzed by NMR techniques (including Diffusion NMR and 2D NOESY), electrospray ionization (ESI) mass spectrometry, and X-ray crystallography. The solid state analysis shows a large change in crystal morphology upon introduction of the endohedral amine groups, caused by deleterious interactions between the amines and the triflate counterions from the coordination process. Combination of different ligands allows analysis of ligand exchange rates via NMR analysis, with half-lives on the order of 3 h, independent of the donor properties of the ligand. Self-sorting behavior is observed, with more electron-rich ligands being favored. The amine-containing and extended complexes are strongly fluorescent, giving quantum yields of up to 83%.     

Application of difference NOE-pumping NMR technique and cold-spray ionization mass spectrometry to identify a ligand binding with a protein receptor.

A difference diffusion-based NMR technique and cold-spray ionization mass spectrometry were employed as a solution-based approach for identifying a ligand binding with a protein receptor. The difference diffusion-based NMR technique, called difference NOE-pumping, can directly detect the ligand interacting with a protein receptor. This technique uses a simple pulse sequence and the diffusion filter can easily be optimized. The cold-spray ionization mass spectrometry (CSI-MS), a variant of electrospray ionization mass spectrometry (ESI-MS) operating at low temperature, has been applied to detect the ligand-receptor complex. The efficiency of these techniques for identifying binding ligands is demonstrated with the human serum albumin (HSA)-drug system.     

NMR-based analysis of protein–ligand interactions

Physiological processes are mainly controlled by intermolecular recognition mechanisms involving protein–protein and protein–ligand (low molecular weight molecules) interactions. One of the most important tools for probing these interactions is high-field solution nuclear magnetic resonance (NMR) through protein-observed and ligand-observed experiments, where the protein receptor or the organic compounds are selectively detected. NMR binding experiments rely on comparison of NMR parameters of the free and bound states of the molecules. Ligand-observed methods are not limited by the protein molecular size and therefore have great applicability for analysing protein–ligand interactions. The use of these NMR techniques has considerably expanded in recent years, both in chemical biology and in drug discovery. We review here three major ligand-observed NMR methods that depend on the nuclear Overhauser effect—transferred nuclear Overhauser effect spectroscopy, saturation transfer difference spectroscopy and water–ligand interactions observed via gradient spectroscopy experiments—with the aim of reporting recent developments and applications for the characterization of protein–ligand complexes, including affinity measurements and structural determination.     

A Series of Diamagnetic Pyridine Monoimine Rhenium Complexes with Different Degrees of Metal‐to‐Ligand Charge Transfer: Correlating 13C NMR Chemical Shifts with Bond Lengths in Redox‐Active Ligands

A set of pyridine monoimine (PMI) rhenium(I) tricarbonyl chlorido complexes with substituents of different steric and electronic properties was synthesized and fully characterized. Spectroscopic (NMR and IR) and single‐crystal X‐ray diffraction analyses of these complexes showed that the redox‐active PMI ligands are neutral and that the overall electronic structure is little affected by the choices of the substituent at the ligand backbone. One‐ and two‐electron reduction products were prepared from selected starting compounds and could also be characterized by multiple spectroscopic methods and X‐ray diffraction. The final product of a one‐electron reduction in THF is a diamagnetic metal–metal‐bonded dimer after loss of the chlorido ligand. Bond lengths in and NMR chemical shifts of the PMI ligand backbone indicate partial electron transfer to the ligand. Two‐electron reduction in THF also leads to the loss of the chlorido ligand and a pentacoordinate complex is obtained. The comparison with reported bond lengths and ¹³C NMR chemical shifts of doubly reduced free pyridine monoaldimine ligands indicates that both redox equivalents in the doubly reduced rhenium complex investigated here are located in the PMI ligand. With diamagnetic complexes varying over three formal reduction stages at the PMI ligand we were, for the first time, able to establish correlations of the ¹³C NMR chemical shifts with the relevant bond lengths in redox‐active ligands over a full redox series.     

Exploring Multi-Subsite Binding Pockets in Proteins: DEEP-STD NMR Fingerprinting and Molecular Dynamics Unveil a Cryptic Subsite at the GM1 Binding Pocket of Cholera Toxin B

Ligand-based NMR techniques to study protein–ligand interactions are potent tools in drug design. Saturation transfer difference (STD) NMR spectroscopy stands out as one of the most versatile techniques, allowing screening of fragments libraries and providing structural information on binding modes. Recently, it has been shown that a multi-frequency STD NMR approach, differential epitope mapping (DEEP)-STD NMR, can provide additional information on the orientation of small ligands within the binding pocket. Here, the approach is extended to a so-called DEEP-STD NMR fingerprinting technique to explore the binding subsites of cholera toxin subunit B (CTB). To that aim, the synthesis of a set of new ligands is presented, which have been subject to a thorough study of their interactions with CTB by weak affinity chromatography (WAC) and NMR spectroscopy. Remarkably, the combination of DEEP-STD NMR fingerprinting and Hamiltonian replica exchange molecular dynamics has proved to be an excellent approach to explore the geometry, flexibility, and ligand occupancy of multi-subsite binding pockets. In the particular case of CTB, it allowed the existence of a hitherto unknown binding subsite adjacent to the GM1 binding pocket to be revealed, paving the way to the design of novel leads for inhibition of this relevant toxin.     

Characterizing mixed phosphonic acid ligand capping on CdSe/ZnS quantum dots using ligand exchange and NMR spectroscopy

The ligand capping of phosphonic acid functionalized CdSe/ZnS core–shell quantum dots (QDs) was investigated with a combination of solution and solid‐state ³¹P nuclear magnetic resonance (NMR) spectroscopy. Two phosphonic acid ligands were used in the synthesis of the QDs, tetradecylphosphonic acid and ethylphosphonic acid. Both alkyl phosphonic acids showed broad liquid and solid‐state ³¹P NMR resonances for the bound ligands, indicative of heterogeneous binding to the QD surface. In order to quantify the two ligand populations on the surface, ligand exchange facilitated by phenylphosphonic acid resulted in the displacement of the ethylphosphonic acid and tetradecylphosphonic acid and allowed for quantification of the free ligands using ³¹P liquid‐state NMR. After washing away the free ligand, two broad resonances were observed in the liquids' ³¹P NMR corresponding to the alkyl and aromatic phosphonic acids. The washed samples were analyzed via solid‐state ³¹P NMR, which confirmed the ligand populations on the surface following the ligand exchange process. Copyright © 2015 John Wiley & Sons, Ltd.     

Molecular-dynamics simulation for the characterization of liquid chromatographic stationary phase: effect of temperature.

The influence of temperature on the surface structure of the octadecylsilica (ODS) bonded phase was investigated with a molecular dynamics (MD) simulation. The MD simulation was applied to a molecular model consisting of three parts: amorphous silica base, dimethyloctadecylsilyl ligands and n-hexane as a mobile phase solvent. More detailed information on the effect of temperature was obtained at the low temperature region than that reported in our previous study. The motion of ODS ligands could be estimated by the mean square displacement (MSD) of the terminal carbon atoms of ODS ligands. The gauche fraction in the ODS ligand conformation can also be estimated to obtain the ligand conformation for each simulation condition in detail. It can be seen that an elevated temperature induced the more bent ligand conformation. The trend has a good agreement to that of the results experimentally observed by using various spectroscopic techniques such as nuclear magnetic resonance (NMR), Fourier-transform infrared spectroscopy (FT-IR), and Raman spectroscopy.     

Gem-diol Generation in Copper and Zinc N-methyl-2-imidazolecarboxaldehyde Complexes: Solid-state NMR,EPR and Single-Crystal X-ray Studies

Novel coordination compounds, mono- ( 1 ) and binuclear copper complexes ( 2 ) and a zinc complex ( 3 ) were synthesized and studied through single-crystal X-ray crystallography, solid-state NMR and EPR techniques to determine the chemical functionalization of the carbonyl group in N-methyl-2-imidazolecarboxaldehyde ligand. Particularly, molecules containing carbonyl groups are versatile ligands that give rise to a wide range of new materials due to the high reactivity of the carbonyl group. However, the chemical identification of the functional group present in these coordination compounds is a challenge because the copper ion affects the NMR signals. In this sense, X-ray crystallography becomes an indispensable tool for the analysis. The imidazole ligands in copper complexes 1 and 2 were found to be the aldehyde and the gem-diol forms, respectively. Furthermore, the gem-diol and carboxylate moieties were detected in the crystal lattice for the zinc complex 3 and studied by solution- and solid-state NMR.     

三种新的铕(III)三元配合物的合成及发光性质研究

合成了吡嗪 [2 ,3 f]并邻菲罗啉及其一系列衍生物 ,作为第二配体 ,并以二苯甲酰甲烷为第一配体 ,合成了 3种新的铕 (III)三元配合物 .通过元素分析、红外光谱、核磁共振光谱确定了它们的组成 ,研究了三种配合物的热稳定性、成膜性能和光致发光性能 (发光强度、荧光量子效率和寿命 ) ,并初步从理论上探讨含有不同基团的第二配体的结构对铕 (III)配合物发光的影响 ,结果表明 :这三种铕 (III)三元配合物均为优良的红色发光材料 ,而且在真空条件下均形成均衡的薄膜 ,这为以这三种铕 (III)配合物作为发光层材料制作有机电致发光器件提供了认识基础     

A New Palladium Complex Containing the Mixture of Carbene and Phosphine Ligands: Synthesis,Crystal Structure and Spectral FT-IR,NMR and UV-Vis Researches

In this paper, a new Pd-based complex that contains N-heterocyclic carbene(NHC) and triphenylphosphine(PPh3) ligands was synthesized. The cis-(NHC)Pd Br_2(PPh3) complex has been prepared by the substitution of 3-chloropyridine ligand in(NHC)PdBr_2(3-chloropyridine) complex with triphenylphosphine. This complex has been characterized by using ~1H NMR, 31 P {~1H} NMR, ~(13)C {~1H} NMR, FTIR, UV-Vis spectroscopy and elemental analysis techniques. The molecular and crystal structures of the cis-(NHC)PdBr_2(PPh3) complex were determined by singlecrystal X-ray diffraction method.     

Formation of a layered framework structure based upon 4-methyl-2,6-bis(methylphosphonic acid) phenol

The trifunctional ligands, [(HO)2P(O)CH2]2C6H2(R)OH, (5-H4)(R = CH3, Br) were prepared in good yield via an Arbusov reaction between P(OEt)3 and the respective 4-R-2,6-bis(chloromethyl)phenols followed by acidic aqueous hydrolysis and they were spectroscopically characterized by IR and NMR techniques. The ligand 5-H4-CH3 readily dissolves lanthanide hydroxide residues and it forms a crystalline complex from aqueous LaCl3 solutions. This complex was characterized by single crystal X-ray diffraction methods and found to adopt a complex 2-D lamellar network in the bc plane. The La(III) inner coordination sphere is seven coordinate formed by oxygen atoms from two water molecules and five phosphonate oxygen atoms from three different ligands. The phenolic oxygen atom is not involved in the ligand binding to La(III).     

Novel Unsymmetric α‐Diimine Nickel(II) Complexes: Suitable Catalysts for Copolymerization Reactions

We established a strategy to synthesize novel unsymmetric 2,3‐diaza‐1,4‐dithiane ligands. Reaction of [Ni(acac)₂] and trityl tetrakis(pentafluorophenyl)borate in the presence of these ligands afforded the corresponding salt‐type complexes. All new compounds were characterized by means of elemental analysis and NMR spectroscopy, and the complexes additionally by mass spectroscopy. NMR spectroscopic experiments on polymers generated by the symmetric ligand/trimethylaluminum catalyst system showed that all products were nearly linear, independent of the polymerization conditions. By contrast, polymers produced by the unsymmetric ligand/trimethylaluminum catalyst system under homopolymerization conditions were branched (15–24 ‰). Additionally, copolymerization experiments with propylene and 1‐hexene afforded copolymers with a branching level of up to 50 ‰.     

Characterization of CdSe quantum dots with bidentate ligands by capillary electrophoresis

The CdSe quantum dots (QDs) with bidentate ligands: a-diimine (NN) and dihydrolipoic acid (DHLA) were synthesized and characterized by UV-Vis, particle size and capillary electrophoretic techniques. Two systems were analyzed: CdSe with one ligand (CdSe/ligand) and CdSe with two different ligands (CdSe//ligand1/ligand2), where ligand = α-diimine or DHLA. Hydrodynamic features of functionalized QDs were characterized by zone capillary electrophoretic (CZE), and particle size techniques and these methods were consistent. It was established that CZE, micellar (MEKC) and microemulsion (MEEKC) modes were suitable for separating charged CdSe QDs and that no peaks were obtained for QDs passivated with electrically neutral ligands. For CdSe QDs with neutral (NN) ligands, a preconcentration method with the use of a micellar plug was introduced for visualizing these QDs. A sharp peak representing neutral QDs was obtained within the zone of micellar plug of a non-ionic surfactant, Here, a ligand character used for CdSe modification and the type of the electrophoretic method applied were the determining factors for the QDs peak visualization. Moreover, examples of visualization of charged and neutral QDs on the same run were presented, and for this purpose, dual mechanism (separation/preconcentration) was proposed.