Project description:The typical linewidths of 1 H NMR spectra of powdered organic solids at 111 kHz magic-angle spinning (MAS) are of the order of a few hundred Hz. While this is remarkable in comparison to the tens of kHz observed in spectra of static samples, it is still the key limit to the use of 1 H in solid-state NMR, especially for complex systems. Here, we demonstrate a novel strategy to further improve the spectral resolution. We show that the anti-z-COSY experiment can be used to reduce the residual line broadening of 1 H NMR spectra of powdered organic solids. Results obtained with the anti-z-COSY sequence at 100 kHz MAS on thymol, β-AspAla, and strychnine show an improvement in resolution of up to a factor of two compared to conventional spectra acquired at the same spinning rate.
Project description:The Bloch-Siegert shift is a phenomenon in NMR spectroscopy and atomic physics in which the observed resonance frequency is changed by the presence of an off-resonance applied field. In NMR, it occurs especially in the context of homonuclear decoupling. Here we develop a practical method for homonuclear decoupling that avoids inducing Bloch-Siegert shifts. This approach enables accurate observation of the resonance frequencies of decoupled nuclear spins. We apply this method to increase the resolution of the HNCA experiment. We also observe a doubling in sensitivity for a 30 kDa protein. We demonstrate the use of band-selective Cβ decoupling to produce amino acid-specific line shapes, which are valuable for assigning resonances to the protein sequence. Finally, we assign the backbone of a 30 kDa protein, Human Carbonic Anhydrase II, using only HNCA experiments acquired with band-selective decoupling schemes, and instrument time of one week.
Project description:Over the years, NMR spectroscopy has become a powerful analytical tool for the identification and quantification of a variety of natural compounds in a broad range of food matrices. Furthermore, NMR can be useful for characterizing food matrices in terms of quality and authenticity, also allowing for the identification of counterfeits. Although NMR requires minimal sample preparation, this technique suffers from low intrinsic sensitivity relative to complementary techniques; thus, the detection of adulterants or markers for authenticity at low concentrations remains challenging. Here, we present a strategy to overcome this limitation by the introduction of a simple band-selective homonuclear decoupling sequence that consists of double irradiation on 1H during NMR signal acquisition. The utility of the proposed method is tested on dihydrosterculic acid (DHSA), one of the cyclopropane fatty acids (CPFAs) shown to be a powerful molecular marker for authentication of milk products. A quantitative description of how the proposed NMR scheme allows sensitivity enhancement yet accurate quantification of DHSA is provided.
Project description:The 1 H NMR analysis of species containing NMR-active heteronuclei can be difficult due to signal overlap caused by the combined effects of homonuclear and heteronuclear scalar (J) couplings. Here, a general pure shift method is presented for obtaining ultra-high resolution 1 H NMR spectra where spectral overlap is drastically reduced by suppressing both homonuclear and heteronuclear J-couplings, giving one single signal per 1 H chemical environment. Its usefulness is demonstrated in the analysis of fluorine- and phosphorus-containing compounds of pharmaceutical and biochemical interest.
Project description:Sensitivity and resolution are the two fundamental obstacles to extending solid-state nuclear magnetic resonance to even larger protein systems. Here, a novel long-observation-window band-selective homonuclear decoupling (LOW BASHD) scheme is introduced that increases resolution up to a factor of 3 and sensitivity up to 1.8 by decoupling backbone alpha-carbon (C(α)) and carbonyl (C') nuclei in U-(13)C-labeled proteins during direct (13)C acquisition. This approach introduces short (<200 μs) pulse breaks into much longer (~8 ms) sampling windows to efficiently refocus the J-coupling interaction during detection while avoiding the deleterious effects on sensitivity inherent in rapid stroboscopic band-selective homonuclear decoupling techniques. A significant advantage of LOW-BASHD detection is that it can be directly incorporated into existing correlation methods, as illustrated here for 2D CACO, NCO, and NCA correlation spectroscopy applied to the β1 immunoglobulin binding domain of protein G and 3D CBCACO correlation spectroscopy applied to the α-subunit of tryptophan synthase.
Project description:2D homonuclear NMR spectroscopy is an essential technique to characterize small and large molecules, such as organic compounds, metabolites, and biomacromolecules at atomic resolution. However, for complex samples 2D homonuclear spectra display poor resolution, making spectral assignment very cumbersome. Here, we propose a new method that exploits the differential T2* relaxation times of individual resonances and resolves the 2D NMR peaks into pseudo-3D spectra, where time is the 3rd dimension. T2* weIghted DEconvolution or TIDE analyzes individual free induction decays (FIDs) and dissects them into sub-FIDs that are transformed into pseudo-3D spectra combining Fourier transformation and covariance NMR. TIDE achieves higher resolution and sensitivity for NMR spectra than classical covariance NMR reducing offset-dependent artifacts. We demonstrate the performance of TIDE for magic angle spinning (MAS) [13C,13C]-DARR NMR spectra of single- and multi-span membrane proteins embedded in lipid bilayers. Since TIDE is applicable to all type of homonuclear correlation experiments for liquid and solid samples, we anticipate that it will be a general method for processing NMR data of biomacromolecules, complex mixtures of metabolites as well as material samples.
Project description:Modern applications of 2D NMR spectroscopy to diagnostic screening, metabolomics, quality control, and other high-throughput applications are often limited by the time-consuming sampling requirements along the indirect time domain t1 . 2D total correlation spectroscopy (TOCSY) provides unique spin connectivity information for the analysis of a large number of compounds in complex mixtures, but standard methods typically require >100 t1 increments for an accurate spectral reconstruction, rendering these experiments ineffective for high-throughput applications. For a complex metabolite mixture it is demonstrated that absolute minimal sampling (AMS), based on direct fitting of resonance frequencies and amplitudes in the time domain, yields an accurate spectral reconstruction of TOCSY spectra using as few as 16 t1 points. This permits the rapid collection of homonuclear 2D NMR experiments at high resolution with measurement times that previously were only the realm of 1D experiments.
Project description:In homonuclear 1H NMR (nuclear magnetic resonance) spectra such as [1H,1H]-NOESY (Nuclear Overhauser Enhancement spectroscopy), which is a historic cornerstone spectrum for biomolecular NMR structural biology, hundreds to thousands of cross peaks are present within a square of approximately 100 ppm2 leading to a lot of signal overlap. Spectral resolution is thus a limiting factor for unambiguous chemical shift assignment and data interpretation for dynamics and structure elucidation. Acquiring the spectra at higher magnetic fields such as at a 1.2 GHz 1H frequency helps to reduce spectral crowding, since resolution scales proportionally to the magnetic field strength. Here, we show that the linewidths of cross peaks in [1H,1H]-NOESY and [1H,1H]-TOCSY spectra can be further reduced by a factor of 2-3 in each dimension by super-resolution spectroscopy. In the indirect dimension a composite exponential-cosine weighted number of scans along the time increments are recorded and digitally smoothened by a window function, while in the direct dimension an exponential-cosine window function is applied. Furthermore, measurement time saving by reduced-acquisition super-resolution (RASR) is introduced. Application to the 20 kDa protein KRAS shows that highly resolved NMR spectra suitable for automated analysis can be acquired within less than 3 hours. The method opens an avenue towards automated chemical shift assignment, dynamics and structure determination of unlabeled small and medium size proteins within 24 hours.
Project description:The resolution of proton solid-state NMR spectra is usually limited by broadening arising from dipolar interactions between spins. Magic-angle spinning alleviates this broadening by inducing coherent averaging. However, even the highest spinning rates experimentally accessible today are not able to completely remove dipolar interactions. Here, we introduce a deep learning approach to determine pure isotropic proton spectra from a two-dimensional set of magic-angle spinning spectra acquired at different spinning rates. Applying the model to 8 organic solids yields high-resolution 1 H solid-state NMR spectra with isotropic linewidths in the 50-400 Hz range.
Project description:Homonuclear two- and multidimensional NMR spectra are standard experiments for the structure determination of small to medium-sized molecules. In the large majority of homonuclear correlated spectra the diagonal contains the most intense peaks. Cross-peaks near the diagonal could overlap with huge tails of diagonal peaks and can therefore be easily overlooked. Here we present a general method for the suppression of peaks along the diagonal in homonuclear correlated spectra. It is based on a spatially selective excitation followed by the suppression of magnetization which has not changed the frequency during the mixing process. In addition to the auto correlation removal, these experiments are also less affected by magnetic field inhomogeneities due to the slice selective excitation, which on the other side leads to a reduced intensity compared to regular homonuclear correlated spectra.