Two- and Three-Dimensional H/C PISEMA Experiments and Their Application to Backbone and Side Chain Sites of Amino Acids and Peptides

Two-dimensional ¹H/¹³C polarization inversion spin exchange at the magic angle experiments were applied to single crystal samples of amino acids to demonstrate their potential utility on oriented samples of peptides and proteins. High resolution is achieved and structural information obtained on backbone and side chain sites from these spectra. A triple-resonance experiment that correlates the ¹H–¹³C_α dipolar coupling frequency with the chemical shift frequencies of the α-carbon, as well as the directly bonded amide ¹⁵N site, is also demonstrated. In this experiment the large ¹H–¹³C_α heteronuclear dipolar interaction provides an independent frequency dimension that significantly improves the resolution among overlapping ¹³C resonances of oriented polypeptides, while simultaneously providing measurements of the ¹³C_α chemical shift, ¹H–¹³C dipolar coupling, and ¹⁵N chemical shift frequencies and angular restraints for backbone structure determination.     

Improved Resolution from Double Constant-Time Evolution of 3D and 4D Triple-Resonance Experiments

Triple-resonance NMR experiments are nearly essential for performing backbone assignments of proteins larger than 15 kDa. Our work extends the double constant-time (2CT) evolution scheme to triple-resonance 3D and 4D experiments. The modifications needed to accomplish 2CT evolution in triple resonance experiments are straight forward, are completely general, and consequently, will yield increased resolution for all out-and-back experiments. We expect that the increased resolution of experiments presented here will be useful in the study of larger proteins (>30 kDa) and in the study of highly helical proteins where¹HN,¹⁵N, and¹³C dimensions are poorly dispersed.     

Three-Dimensional C Shift/H–N Coupling/N Shift Solid-State NMR Correlation Spectroscopy

Triple-resonance experiments capable of correlating directly bonded and proximate carbon and nitrogen backbone sites of uniformly ¹³C- and ¹⁵N-labeled peptides in stationary oriented samples are described. The pulse sequences integrate cross-polarization from ¹H to ¹³C and from ¹³C to ¹⁵N with flip-flop (phase and frequency switched) Lee–Goldburg irradiation for both ¹³C homonuclear decoupling and ¹H–¹⁵N spin exchange at the magic angle. Because heteronuclear decoupling is applied throughout, the three-dimensional pulse sequence yields ¹³C shift/¹H–¹⁵N coupling/¹⁵N shift correlation spectra with single-line resonances in all three frequency dimensions. Not only do the three-dimensional spectra correlate ¹³C and ¹⁵N resonances, they are well resolved due to the three independent frequency dimensions, and they can provide up to four orientationally dependent frequencies as input for structure determination. These experiments have the potential to make sequential backbone resonance assignments in uniformly ¹³C- and ¹⁵N-labeled proteins.     

Multiplicity-Selective Coherence Transfer Steps for the Design of Amino Acid-Selective Experiments-A Triple-Resonance Experiment Selective for Asn and Gln

A multiplicity-selective coherence transfer step is discussed, that can replace the normal INEPT transfer in triple-resonance experiments. Depending on the pulse sequence in which they are implemented, amino acid-selective experiments will be created. Two experiments selective for Asn and Gln are proposed.     

钠分子里德堡能级的无多普勒加宽光光三共振光谱以及能级宽度和谱线位移

首次报道了钠分子一系列里德堡能级的无多普勒加宽光光三共振（ＤＦ－ＯＯＴＲ）激发荧光光谱，测量了其谱线宽度和其谱线位置随温度、外加气体压力的变化情况。结果表明，钠分子里德堡能级（ｎ＝５～８）的自然宽度（含均匀加宽）在６０～１３０ＭＨｚ之间，其大小主要与主量子数ｎ有关。随钠原子浓度增大（温度升高），或外加Ａｒ气压力增大时，钢分子各个里德堡能级谱线均出现红移，数量在１００ＭＨｚ／τ之内，但位移速率大小不一。     

A set of triple-resonance nuclear magnetic resonance experiments for structural characterization of organophosphorus compounds in mixture samples

The ¹H, ¹³C correlation NMR spectroscopy utilizes J_CH couplings in molecules, and provides important structural information from small organic molecules in the form of carbon chemical shifts and carbon-proton connectivities. The full potential of the ¹H, ¹³C correlation NMR spectroscopy has not been realized in the Chemical Weapons Convention (CWC) related verification analyses due to the sample matrix, which usually contains a high amount of non-related compounds obscuring the correlations of the relevant compounds. Here, the results of the application of ¹H, ¹³C, ³¹P triple-resonance NMR spectroscopy in characterization of OP compounds related to the CWC are presented. With a set of two-dimensional triple-resonance experiments the J_HP, J_CH and J_PC couplings are utilized to map the connectivities of the atoms in OP compounds and to extract the carbon chemical shift information. With the use of the proposed pulse sequences the correlations from the OP compounds can be recorded without significant artifacts from the non-OP compound impurities in the sample. Further selectivity of the observed correlations is achieved with the application of phosphorus band-selective pulse in the pulse sequences to assist the analysis of multiple OP compounds in mixture samples. The use of the triple-resonance experiments in the analysis of a complex sample is shown with a test mixture containing typical scheduled OP compounds, including the characteristic degradation products of nerve agents sarin, soman, and VX. The viability of the approach in verification analysis is demonstrated in the analysis of the 30th OPCW Proficiency Test sample.