Multidimensional solid-state NMR spectroscopy of plant cell walls

Plant biomass has become an important source of bio-renewable energy in modern society. The molecular structure of plant cell walls is difficult to characterize by most atomic-resolution techniques due to the insoluble and disordered nature of the cell wall. Solid-state NMR (SSNMR) spectroscopy is uniquely suited for studying native hydrated plant cell walls at the molecular level with chemical resolution. Significant progress has been made in the last five years to elucidate the molecular structures and interactions of cellulose and matrix polysaccharides in plant cell walls. These studies have focused on primary cell walls of growing plants in both the dicotyledonous and grass families, as represented by the model plants Arabidopsis thaliana, Brachypodium distachyon, and Zea mays. To date, these SSNMR results have shown that 1) cellulose, hemicellulose, and pectins form a single network in the primary cell wall; 2) in dicot cell walls, the protein expansin targets the hemicellulose-enriched region of the cellulose microfibril for its wall-loosening function; and 3) primary wall cellulose has polymorphic structures that are distinct from the microbial cellulose structures. This article summarizes these key findings, and points out future directions of investigation to advance our fundamental understanding of plant cell wall structure and function.     

High-resolution solid-state NMR spectroscopy as a tool for investigation of enantioselective inclusion complexation

In this paper, we showed the application of solid state-NMR (SS NMR) spectroscopy in structural studies of chiral compounds employing sample of (E)-1-diphenylphosphinoylpent-3-en-2-ol 1 as a model compound. Racemate of 1 was fully characterized by NMR techniques (both in liquid and solid phase) and X-ray crystallography. Theoretical calculations employing the GIAO approach were used to explain the influence of hydrogen bonding on 31P NMR shielding parameter in racemate. Enantioselective inclusion complexation (EIC) method with TADDOL as host molecule was applied to separate of enantiomers. The formation of host-guest complex and decomplexation procedure was monitored by means of the SS NMR. The liquid-state NMR, due to similarity of 13C and 31P spectral parameters was not able to distinguish racemate from enantiomer. In the solid phase, owing to distinction of hydrogen bonding and molecular packing in the crystal lattice, racemate and enantiomers were easy recognized by NMR spectroscopy.     

High-resolution solid-state NMR spectroscopy of protons with homonuclear dipolar decoupling schemes under magic-angle spinning

High-resolution NMR spectroscopy of ¹H spins in the solid state is normally rendered difficult due to the strong homonuclear ¹H–¹H dipolar couplings. Even under very high-speed magic-angle spinning (MAS) at ca. 60–70 kHz, these couplings are not completely removed. An appropriate radiofrequency pulse scheme is required to average out the homonuclear dipolar interactions in combination with MAS to get high-resolution ¹H NMR spectrum in solid state. Several schemes have been introduced in the recent past with a variety of applications also envisaged. Development of some of these schemes has been made possible with a clear understanding of the underlying spin physics based on bimodal Floquet theory. The utility of these high-resolution pulse schemes in combination with MAS has been demonstrated for spinning speeds of 10–65 kHz in a range of ¹H Larmor frequencies from 300 to 800 MHz.     

Pure-exchange solid-state NMR

Three exchange nuclear magnetic resonance (NMR) techniques are presented that yield (13)C NMR spectra exclusively of slowly reorienting segments, suppressing the often dominant signals of immobile components. The first technique eliminates the diagonal ridge that usually dominates two-dimensional (2D) exchange NMR spectra and that makes it hard to detect the broad and low off-diagonal exchange patterns. A modulation of the 2D exchange spectrum by the sine-square of a factor which is proportional to the difference between evolution and detection frequencies is generated by fixed additional evolution and detection periods of duration tau, yielding a 2D pure-exchange (PUREX) spectrum. Smooth off-diagonal intensity is obtained by systematically incrementing tau and summing up the resulting spectra. The related second technique yields a static one-dimensional (1D) spectrum selectively of the exchanging site(s), which can thus be identified. Efficient detection of previously almost unobservable slow motions in a semicrystalline polymer is demonstrated. The third approach, a 1D pure-exchange experiment under magic-angle spinning, is an extension of the exchange-induced sideband (EIS) method. A TOSS (total suppression of sidebands) spectrum obtained after the same number of pulses and delays, with a simple swap of z periods, is subtracted from the EIS spectrum, leaving only the exchange-induced sidebands and a strong, easily detected centerband of the mobile site(s).     

High-resolution solid-state NMR of anisotropically mobile molecules under very low-power H decoupling and moderate magic-angle spinning

We show that for observing high-resolution heteronuclear NMR spectra of anisotropically mobile systems with order parameters less than 0.25, moderate magic-angle spinning (MAS) rates of 11 kHz combined with ¹H decoupling at 1–2 kHz are sufficient. Broadband decoupling at this low ¹H nutation frequency is achieved by composite pulse sequences such as WALTZ-16. We demonstrate this moderate MAS low-power decoupling technique on hydrated POPC lipid membranes, and show that 1 kHz ¹H decoupling yields spectra with the same resolution and sensitivity as spectra measured under 50 kHz ¹H decoupling when the same acquisition times (50 ms) are used, but the low-power decoupled spectra give higher resolution and sensitivity when longer acquisition times (>150 ms) are used, which are not possible with high-power decoupling. The limits of validity of this approach are explored for a range of spinning rates and molecular mobilities using more rigid membrane systems such as POPC/cholesterol mixed bilayers. Finally, we show ¹⁵N and ¹³C spectra of a uniaxially diffusing membrane peptide assembly, the influenza A M2 transmembrane domain, under 11 kHz MAS and 2 kHz ¹H decoupling. The peptide ¹⁵N and ¹³C intensities at low-power decoupling are 70–80% of the high-power decoupled intensities. Therefore, it is possible to study anisotropically mobile lipids and membrane peptides using liquid-state NMR equipment, relatively large rotors, and moderate MAS frequencies.     

High-resolution solid-state 13C NMR studies of poly[(R)-3-hydroxybutyric] acid

Solid poly[(R)-3-hydroxybutyric] acid was examined by high-resolution ¹³C MAS NMR, differential scanning calorimetry and infrared spectroscopy. The ¹³C methyl group resonance consists of three components: the rigid amorphous phase, the crystalline phase and the mobile amorphous phase. Spectral deconvolution, using the Lorentz function, reveals the relative amounts as 62% crystalline and 38% amorphous at 333 K, and 42% crystalline, 40% pure amorphous and 18% β-orthorhombic at 413 K. NMR indicates a large difference in molecular mobility between the crystalline and amorphous regions of the sample. Infrared spectroscopy shows that the stretching at 1725 cm⁻¹ (characteristic of the -form) comes from the crystalline region, and the bands at 1744 cm⁻¹ (characteristic of the β-form) and 1800 cm⁻¹ come from the amorphous region.     

一种新的HETISINE型二萜生物碱结构的NMR研究

从毛莨科乌头属黄花乌头的根中分得一个化合物,经过测定,它是hetisine型二萜生物碱,其结构为13-乙酰基-14-羟基-2-丙酰基亥替钦,为一新化合物.本文报道选择性远程DEPT技术在该化合物的结构和NMR信号归属研究中的应用.     

A solid-state NMR index of helical membrane protein structure and topology

The secondary structure and topology of membrane proteins can be described by inspection of two-dimensional (1)H-(15)N dipolar coupling/(15)N chemical shift polarization inversion spin exchange at the magic angle spectra obtained from uniformly (15)N-labeled samples in oriented bilayers. The characteristic wheel-like patterns of resonances observed in these spectra reflect helical wheel projections of residues in both transmembrane and in-plane helices and hence provide direct indices of the secondary structure and topology of membrane proteins in phospholipid bilayers. We refer to these patterns as PISA (polarity index slant angle) wheels. The transmembrane helix of the M2 peptide corresponding to the pore-lining segment of the acetylcholine receptor and the membrane surface helix of the antibiotic peptide magainin are used as examples.     

PVPh/PEO共混物动力学演化过程的NMR研究

聚合物共混物中链段的慢取向运动与其玻璃化转变行为和宏观力学性质密切关联,而基于化学位移各向异性重聚的~(13)C CODEX(centerband-only detection of exchange)固体核磁共振(SSNMR)技术能够有效表征共混物中链段的慢取向运动.该文利用~(13)C CODEX NMR技术详细研究了相容性聚合物共混物聚乙烯基苯酚/聚氧乙烯(PVPh/PEO)中的刚性组分PVPh在较宽温度范围内的慢取向运动特性与玻璃化转变过程的关联.研究表明,在玻璃化转变起始温度以下,PVPh主链的分子运动被冻结,而侧基存在b-松弛的慢取向运动;在玻璃化转变起始温度附近,PVPh主链具有明显的慢取向运动,而且主链和侧基是一种协同的分子运动.该文利用NMR技术揭示了共混物中的玻璃化转变起止温度分别对应于高分子主链慢取向运动CODEX信号的开始和极大值处的温度.     

High-resolution NMR studies of nifedipine

The paper reports results of¹H and¹³C high-resolution nuclear magnetic resonance studies of nifedipine and signal assignment to particular atoms. High-resolution¹H NMR spectra with no signals attributed to protons in weak fields (δ= 10−12 ppm) imply that H7 atom is strongly connected to the ring and is not involved in the formation of hydrogen bonds.     

Structure determination of aligned systems by solid-state NMR magic angle spinning methods

Single crystal rotational echo double resonance (REDOR) experiments can be used to determine the three-dimensional orientation of heteronuclear bond vectors in an amino acid, as well as the crystal's orientation relative to the rotor fixed frame (RFF). We also demonstrate that for samples uniaxially aligned along the rotor axis, the polar tilt angle of a bond vector relative to the RFF can be measured by use of an analytical expression that describes the REDOR curve for that system. These bond orientations were verified by X-ray indexing of the single crystal sample, and were shown to be as accurate as +/- 1 degrees .     

Spin-state selection in solid-state NMR

Spin-state selection in solid-state NMR is demonstrated, using similar pulse sequences as used in liquid-state NMR. The different transitions of all three carbon resonances in fully 13C-labeled L-alanine are separated in different spectra. By selecting spin-states, the contribution of the J-coupling to the linewidth is removed, leading to a considerable enhancement in resolution. The spin-state-selective technique is demonstrated for magic-angle spinning frequencies from 6 to 35kHz. Other experimental conditions affecting the sensitivity of the experiments are discussed. Sensitivity losses due to the introduction of the spin-state-selective filter are shown to be acceptable. Finally, spin-state selection was used to experimentally confirm the differential broadening expected for the two transitions of the CH3 resonance.     

一种新的DENUDATINE型C20二萜生物碱结构的NMR研究

从内蒙西伯利亚乌头中分离得一个新的C₂₀二萜生物碱,采用选择性远程DEPT,同核和异核二维NMR技术相结合进行了研究,其结构确定为Lepenine的C-11基向立体异构体,定名为11a-hydroxylapenine.结果表明,选择性远程DEPTNMR技术对于连接这类化合物中被季碳和杂原子分割的NMR自旋体系,测定基本骨架和确定信号归属都有独到之处.     

A solid-state NMR investigation of the structure of mesoporous silica nanoparticle supported rhodium catalysts

A detailed study of the chemical structure of mesoporous silica catalysts containing rhodium ligands and nanoparticles (RhP-MSN) was carried out by multi-dimensional solid-state NMR techniques. The degree of functionalization of the rhodium–phosphinosilyl complex to the surface of the RhP-MSN channels was determined by ²⁹Si NMR experiments. The structural assignments of the rhodium–phosphinosilyl complex were unambiguously determined by employing the novel, indirectly detected heteronuclear correlation (¹³C–¹H and ³¹P–¹H idHETCOR) techniques, which indicated that oxidation of the attached phosphinosilyl groups and detachment of Rh was enhanced upon syngas conversion.     

Operator-based Floquet theory in solid-state NMR

This article reviews the application of operator-based Floquet theory in solid-state NMR. Basic expressions for calculating effective Hamiltonians based on van Vleck perturbation theory are reviewed for problems with a single frequency or multiple incommensurate frequencies. Such a treatment allows calculation of effective Hamiltonians for resonant and non-resonant problems. Examples from literature are given for single-mode to triple-mode Floquet problems, covering a wide range of applications in solid-state NMR under magic-angle spinning and radio-frequency irradiation of a single nucleus or multiple nuclei.     

一种新的DENUDATINE型C20二萜生物碱结构的NMR研究

从内蒙西伯利亚乌头中分离得一个新的C20二萜生物碱,采用选择性远程DEPT,同核和异核二维NMR技术相结合进行了研究,其结构确定为Lepenine的C－11基向立体异构体,定名为11a－hydroxylapenine．结果表明,选择性远程DEPTNMR技术对于连接这类化合物中被季碳和杂原子分割的NMR自旋体系,测定基本骨架和确定信号归属都有独到之处．     

Advances in solid-state NMR of membrane proteins

Solid-state nuclear magnetic resonance (SSNMR) is an NMR spectroscopy applied to condensed-phase systems, including membrane proteins. Membrane protein fold and function are dependent upon interactions with surrounding bilayer components. Structural and functional analyses are thus challenging, and new approaches are needed to better characterise these systems. SSNMR is uniquely suited to the examination of membrane proteins in native environments, and has the capabilities to elucidate complex protein mechanisms and structures. Notable research implementing SSNMR is aimed at developing new strategies and technology to efficiently target membrane proteins within synthetic and biological membranes. Significant advances have been made: observation of protein function in native environments, emergence of in situ methods to examine integral proteins within natural membranes, sensitivity enhancement techniques and cutting-edge structure determination methods. We present how these advances are applied to answer outstanding questions in structural biology. Experiments have shown consistent results for protein investigations in biological membranes and synthetic lipid compositions, indicating that SSNMR is an innovative and direct approach for the study of these systems.