The Effect of Trifluoroacetic Acid on Molecular Weight Determination of Polyesters:An in Situ NMR Investigation

Due to the poor solubility of aromatic polyesters in common organic solvents,trifluoroacetic acid is usually used as a co-solvent to increase their solubility for characterizations.However,only few studies have reported the side reactions induced by it.We present here the application of in situ 1H-NMR techniques to explore its effect on the hydroxyl end-groups,which are usually used for the molecular weight determination of polyesters by end-group estimation method.Using bis(2-hydroxyethyl) terephthalate (BHET) as model compound,1H quantitative NMR results show the peak integration of hydroxyethyl end-groups decreased with time via a pseudo-first-order kinetics in d-trifluoroacetic acid/d-chloroform mixture solvent (1∶10,V∶V).This is due to the esterification of hydroxyethyl groups with trifluoroacetic acid,revealed by the 1H-13C gradient-enhanced heteronuclear multiple bond correlation (gHMBC) spectrum.The mixtures of dimethyl terephthalate and BHET with different molar ratios were used to represent poly(ethylene terephthalate) (PET) with different degrees of polymerization,and the effect of trifluoroacetic acid on the estimation of hydroxyethyl groups and subsequent molecular weight determination of polyesters was studied.Our results show that if a relative error of 5％ is allowed,the NMR measurements must be finished within 1.3 h of solution preparation at 25 ℃ in the mixture solvent.The results were confirmed in PET sample,while in poly(ethylene adipate),the obtained esterifaction constant is faster that those in aromatic system.The results can be applied to other polymer systems with alcohol functionalized groups,and used as a guideline for the characterization of polyesters and polyethers by end-group estimation method.     

疏水缔合聚合物分子量分布曲线的测定

在体积百分浓度为50%1,3-丙二醇、0.2 mol/L Na Cl和水溶剂条件下,消除疏水缔合聚合物(HAWSP)溶液中疏水缔合作用,屏蔽溶液中的聚电解质效应,使HAWSP分子在稀溶液中处于单分子分散状态.然后利用膜孔径分离原理,选择不同孔径的微孔滤膜,用微孔滤膜流动实验装置对疏水缔合聚合物进行分级,将不同分子量的聚合物分离开来.用二次方程拟合滤出液质量-过滤时间关系曲线得到各级分聚合物溶液的质量,以分光光度法测定各级分聚合物溶液的浓度,根据质量和浓度计算得到各级分的累积百分含量.结合静态光散射和毛细管法标定了疏水缔合聚合物Mark-Houwink方程[η]=0.182M~(0.586),用于准确测定疏水缔合聚合物各个级分的分子量.选择四参数方程曲线,根据各级分的分子量M和累积百分含量W,得到分子量的分布曲线.与动态光散射分析进行比较的结果表明,两种方法测试结果一致.     

Magnetically Induced Alignment of Natural Products for Stereochemical Structure Determination via NMR

Anisotropic NMR has gained increasing popularity to determine the structure and specifically the configuration of small, flexible, non‐crystallizable molecules. However, it suffers from the necessity to dissolve the analyte in special media such as liquid crystals or polymer gels. Generally, small degrees of alignment are also caused by an anisotropic magnetic susceptibility of the molecule, for example, induced by aromatic moieties. For this mechanism, the alignment can be predicted via density functional theory. Here we show that both residual dipolar couplings and residual chemical shift anisotropies can be acquired from natural products without special sample preparation using magnetically induced alignment. On the two examples of the novel natural product gymnochrome G and the alkaloid strychnine, these data, together with the predicted alignment, yield the correct configuration with high certainty.     

略论分子量测定在大分子概念形成中的作用

本文介绍了高分子概念的起源,对大分子的早期认识,分子量测定方法的发展和大分子概念的确立过程,讨论了分子量测定方法在大分子概念形成中的作用。     

Combinatorial Synthesis and Multidimensional NMR Spectroscopy: An Approach to Understanding Protein–Ligand Interactions

Combinatorial chemistry is a laboratory emulation of natural recombination and selection processes. Strategies in this developing discipline involve the generation of diverse, molecular libraries through combinatorial synthesis and the selection of compounds that possess a desired property. Such approaches can facilitate the identification of ligands that bind to biological receptors, promoting our chemical understanding of cellular processes. This article illustrates that the coupling of combinatorial synthesis, multidimensional NMR spectroscopy, and biochemical methods has enhanced our understanding of a protein receptor used commonly in signal transduction, the Src Homology 3 (SH3) domain. This novel approach to studying molecular recognition has revealed a set of rules that govern SH3–ligand interactions, allowing models of receptor–ligand complexes to be constructed with only a knowledge of the polypeptide sequences. Combining combinatorial synthesis with structural methods provides a powerful new approach to understanding how proteins bind their ligands in general.     

1H NMR Spectroscopy as an Effective Method for Predicting Molecular Weight of Polyaminoamine Dendrimers and Their Derivatives

We established two formulas to predict molecular weight of polyaminoamine dendrimers and their alkylated derivatives, based on the theoretical number of protons at specific positions in the dendrimers and the true value of the integral values of these protons appearing in proton nuclear magnetic resonance spectra. Calculated results indicated that molecular weight of the dendrimers is approximately equal to results from mass spectrometry. Degrees of alkylation were easily calculated for each dendrimer-alkylated derivative. According to the obtained results, we confirm that the use of the proton spectra can be an effective method to predict molecular weight of dendrimers.     

High‐Resolution NMR Determination of the Dynamic Structure of Membrane Proteins

¹⁵N spin‐relaxation rates are demonstrated to provide critical information about the long‐range structure and internal motions of membrane proteins. Combined with an improved calculation method, the relaxation‐rate‐derived structure of the 283‐residue human voltage‐dependent anion channel revealed an anisotropically shaped barrel with a rigidly attached N‐terminal helix. Our study thus establishes an NMR spectroscopic approach to determine the structure and dynamics of mammalian membrane proteins at high accuracy and resolution.     

Structure Determination of Cellulose Esters via Subsequent Functionalization and NMR Spectroscopy

New paths for the fast and reliable analysis of cellulose esters (CE) via subsequent functionalization and ¹H NMR spectroscopy were studied. Perpropionylation of the CE is an inexpensive and efficient method. For cellulose diacetates used as representative ester well resolved ¹H NMR spectra were obtained, which can be used for the calculation of the over all degree of substitution (DS) and the partial DS values at position 2, 3, and 6. No transesterification occurs during the subsequent acylation and a standard deviation of S² = 1.32 x 10⁻⁴ was found for a series of experiments. In case of more complex ester structures especially with extended aliphatic moieties per-4-nitrobenzoylation need to be applied prior to NMR measurements. The spectra obtained can be completely assigned and applied for the calculation of DS values.     

NMR和分子力学法研究溶液中有机分子结构

在分子力学能量优化项中加入由ＮＭＲ实验得到的几何参数二面角（相应于~３Ｊ_（ＨＨ）耦合常数）及质子间距所构造的约束函数，使分子力学计算得到的能量极小点的分子构象，其耦合常数（~３Ｊ_（ＨＨ））及质子间距同ＮＭＲ实验结果相符合。文中以（±）－３－（４＇－甲苯基）－１－氮杂双环［２．２．２］－３－辛醇为例，采用自编的ＮＭＲ参数约束的分子力学计算程序ＭＭ２ＮＪ对其构象进行了计算。所得结果与Ｘ射线衍射法比较，有较好的一致性     

Non‐Uniform Sampling and J‐UNIO Automation for Efficient Protein NMR Structure Determination

High‐resolution structure determination of small proteins in solution is one of the big assets of NMR spectroscopy in structural biology. Improvements in the efficiency of NMR structure determination by advances in NMR experiments and automation of data handling therefore attracts continued interest. Here, non‐uniform sampling (NUS) of 3D heteronuclear‐resolved [¹H,¹H]‐NOESY data yielded two‐ to three‐fold savings of instrument time for structure determinations of soluble proteins. With the 152‐residue protein NP_372339.1 from Staphylococcus aureus and the 71‐residue protein NP_346341.1 from Streptococcus pneumonia we show that high‐quality structures can be obtained with NUS NMR data, which are equally well amenable to robust automated analysis as the corresponding uniformly sampled data.     

Two-Dimensional Solid-State NMR Spectroscopy: New Possibilities for the Investigation of the Structure and Dynamics of Solid Polymers [New Analytical Methods (38)]

NMR spectroscopy is an effective method not only for examining liquid samples but also for characterizing molecular sturcture, order and dynamics in amorphous and ordered solids. Recent developments in the area of solid-state NMR spectroscopy span from model-dependent studies of conventional one-dimensional spectra to the more definitive two-dimensional (2D) spectra which provide more specific information. For example, with 2D-NMR spectroscopy it is possible to determine the orientational distribution functions of molecular segments in drawn polymers and to distinguish different mechanisms of complex molecular motions. Following an introduction to basic NMR spectroscopy, an overview of the current state-of-the-art of 2D methods in solid-state NMR spectroscopy is presented and demonstrated with selected examples.     

MALDI／MS快速测定蛋清中溶菌酶的相对分子质量

本文介绍了一种用基本辅助激光解吸电离质谱法快速测定蛋清中溶菌酶相对分子质量的方法。与传统的蛋白质相对分子质量测定相比，ＭＡＬＤＩ／ＭＳ法具有灵敏度高、精度高、易操作和分析时间短的优点，包括样品处理在内，一次分析不超过３０ｍｉｎ。     

Determination of Molecular Weight of PDPS Modified Copolymers

Molecular weight (MW) determinations of polydiphenylsiloxane‐co‐polydimethylsiloxane (PDPS/PDMS), polydiphenylsiloxane‐co‐polymethylphenylsiloxane (PDPS/PMPS) and polydiphenylsiloxane‐co‐polymethyl(3,3,3‐trifluoropropyl)siloxane (PDPS/PMFPS) statistical copolymers by gel permeation chromatography (GPC), GPC coupled differential viscometer (GPC‐DV) and light scattering (LS) techniques have been compared and discussed. The MW obtained by GPC‐DV in the PDPS/PDMS and the PDPS/PMPS series agreed with that obtained by LS very well. In PDPS/PDMS copolymers, it was found that the MW obtained by GPC was much lower than that obtained by GPC‐DV and LS, as Ph₂SiO) content is higher than 50 mol%. In PDPS/PMFPS copolymers, the MW obtained by GPC was far different from that obtained by GPC‐DV and the deviation decreased with increasing Ph₂SiO) mol%. The α values of the copolymers can be explained by the structure of the polymer in tetrahydrofuran (THF). Based on the relatively soluble copolymers, not only the differential refractive index increments (dn/dc) of PMPS, PDMS and PMFPS homopolymers but also that of the PDPS homopolymer in THF could be calculated by their corresponding copolymers.     

Structural Aspects of the O-glycosylation Linkage in Glycopeptides via MD Simulations and Comparison with NMR Experiments

A powerful conformational searching and enhanced sampling simulation method, and unbiased molecular dynamics simulations have been used along with NMR spectroscopic observables to provide a detailed structural view of O-glycosylation. For four model systems, the force-field parameters can accurately predict experimental NMR observables (J couplings and NOE's). This enables us to derive conclusions based on the generated ensembles, in which O-glycosylation affects the peptide backbone conformation by forcing it towards to an extended conformation. An exception is described for β-GalNAc-Thr where the α content is increased and stabilized via hydrogen bonding between the sugar and the peptide backbone, which was not observed in the rest of the studied systems. These observations might offer an explanation for the evolutionary preference of α-linked GalNAc glycosylation instead of a β link.     

NMR Spectroscopic Characterization of the C-Mannose Conformation in a Thrombospondin Repeat Using a Selective Labeling Approach

Despite the great interest in glycoproteins, structural information reporting on conformation and dynamics of the sugar moieties are limited. We present a new biochemical method to express proteins with glycans that are selectively labeled with NMR-active nuclei. We report on the incorporation of ¹³C-labeled mannose in the C-mannosylated UNC-5 thrombospondin repeat. The conformational landscape of the C-mannose sugar puckers attached to tryptophan residues of UNC-5 is characterized by interconversion between the canonical ¹C₄ state and the B₀₃ / ¹S₃ state. This flexibility may be essential for protein folding and stabilization. We foresee that this versatile tool to produce proteins with selectively labeled C-mannose can be applied and adjusted to other systems and modifications and potentially paves a way to advance glycoprotein research by unravelling the dynamical and conformational properties of glycan structures and their interactions.     

Structure Determination of Binuclear Triple-Decker Phthalocyaninato Complexes by NMR via Paramagnetic Shifts Analysis Using Symmetry Peculiarities

The detailed knowledge about the structure of multinuclear paramagnetic lanthanide complexes for the targeted design of these compounds with special magnetic, sensory, optical and electronic properties is a very important task. At the same time, establishing the structure of such multinuclear paramagnetic lanthanide complexes in solution, using NMR is a difficult task, since several paramagnetic centers act simultaneously on the resulting chemical shift of a particular nucleus. In this paper, we have demonstrated the possibility of molecular structure determination in solution on the example of binuclear triple-decker lanthanide(III) complexes with tetra-15-crown-5-phthalocyanine Ln₂[(15C5)₄Pc]₃ {where Ln = Tb (1) and Dy (2)} by quantitative analysis of the pseudo-contact lanthanide-induced shifts (LIS). The symmetry of complexes was used for the simplification of the calculation of pseudo-contact shifts on the base of the expression for the magnetic susceptibility tensor in the arbitrary oriented magnetic axis system. Good agreement between the calculated and experimental shifts in the ¹H NMR spectra indicates the similarity of the structure for the complexes 1 and 2 in solution of CDCl₃ and the structure in the crystalline phase, found from the data of the X-ray structural study of the similar complex Lu₂[(15C5)₄Pc]₃. The described approach can be useful for LIS analysis of other polynuclear symmetric lanthanide complexes.     

Reading Polymers: Sequencing of Natural and Synthetic Macromolecules

The sequencing of biopolymers such as proteins and DNA is among the most significant scientific achievements of the 20th century. Indeed, modern chemical methods for sequence analysis allow reading and understanding the codes of life. Thus, sequencing methods currently play a major role in applications as diverse as genomics, gene therapy, biotechnology, and data storage. However, in terms of fundamental science, sequencing is not really a question of molecular biology but rather a more general topic in macromolecular chemistry. Broadly speaking, it can be defined as the analysis of comonomer sequences in copolymers. However, relatively different approaches have been used in the past to study monomer sequences in biological and manmade polymers. Yet, these “cultural” differences are slowly fading away with the recent development of synthetic sequence‐controlled polymers. In this context, the aim of this Minireview is to present an overview of the tools that are currently available for sequence analysis in macromolecular science.     

Studies Related to Norway Spruce Galactoglucomannans: Chemical Synthesis,Conformation Analysis,NMR Spectroscopic Characterization,and Molecular Recognition of Model Compounds

Galactoglucomannan (GGM) is a polysaccharide mainly consisting of mannose, glucose, and galactose. GGM is the most abundant hemicellulose in the Norway spruce (Picea abies), but is also found in the cell wall of flax seeds, tobacco plants, and kiwifruit. Although several applications for GGM polysaccharides have been developed in pulp and paper manufacturing and the food and medical industries, attempts to synthesize and study distinct fragments of this polysaccharide have not been reported previously. Herein, the synthesis of one of the core trisaccharide units of GGM together with a less‐abundant tetrasaccharide fragment is described. In addition, detailed NMR spectroscopic characterization of the model compounds, comparison of the spectral data with natural GGM, investigation of the acetyl‐group migration phenomena that takes place in the polysaccharide by using small model compounds, and a binding study between the tetrasaccharide model fragment and a galactose‐binding protein (the toxin viscumin) are reported.     

Structure of a cyclic peptide with a catalytic triad, determined by computer simulation and NMR spectroscopy

Summary We report the design of a cyclic, eight-residue peptide that possesses the catalytic triad residues of the serine proteases. A manually built model has been relaxed by 0.3 ns of molecular dynamics simulation at room temperature, during which no major changes occurred in the peptide. The molecule has been synthesised and purified. Two-dimensional NMR spectroscopy provided 35 distance and 7 torsion angle constraints, which were used to determine the three-dimensional structure. The experimental conformation agrees with the predicted one at the -turn, but deviates in the arrangement of the disulphide bridge that closes the backbone to a ring. A 1.2 ns simulation at 600 K provided extended sampling of conformation space. The disulphide bridge reoriented into the experimental arrangement, producing a minimum backbone rmsd from the experimental conformation of 0.8 . At a later stage in the simulation, a transition at Ser³ produced more pronounced high-temperature behaviour. The peptide hydrolyses p-nitrophenyl acetate about nine times faster than free histidine.