New Article in Journal of Biomolecular NMR
Multidimensional NMR can provide unmatched spectral resolution, which is crucial when dealing with samples of biological macromolecules. The resolution, however, comes at the high price of long experimental time. Non-uniform sampling (NUS) of the evolution time domain allows to suppress this limitation by sampling only a small fraction of the data, but requires sophisticated algorithms to reconstruct omitted data points. A significant group of such algorithms known as compressed sensing (CS) is based on the assumption of sparsity of a reconstructed spectrum. Several papers on the application of CS in multidimensional NMR have been published in the last years, and the developed methods have been implemented in most spectral processing software. However, the publications rarely show the cases when NUS reconstruction does not work perfectly or explain how to solve the problem. On the other hand, every-day users of NUS develop their rules-of-thumb, which help to set up the processing in an optimal way, but often without a deeper insight. In this paper, we discuss several sources of problems faced in CS reconstructions: low sampling level, missassumption of spectral sparsity, wrong stopping criterion and attempts to extrapolate the signal too much. As an appendix, we provide MATLAB codes of several CS algorithms used in NMR. We hope that this work will explain the mechanism of NUS reconstructions and help readers to set up acquisition and processing parameters. Also, we believe that it might be helpful for algorithm developers.
New Article in ChemComm
The dangerously high power levels required for fast ASAP-HSQC and ASAP-HMQC experiments are mitigated by employing EXACT NMR. The utility of this technique is demonstrated by application of the EXACT ASAP-HSQC to chemical reaction monitoring, accelerating data acquisition by up to 2 orders of magnitude – providing evidence of it’s value in fast quantitative NMR processes.
New Article in Journal of Peptide Science
The impact of β-azido(or 1-piperidinyl)methylamino acids in position 2 or 3 on biological activity and conformation of dermorphin analogues
Maciej Maciejczyk, Anika Lasota, Oliwia Frączak, Piotr Kosson, Aleksandra Misicka, Michal Nowakowski, Andrzej Ejchart and Aleksandra Olma
The synthesis of new dermorphin analogues is described. The (R)-alanine or phenylalanine residues of natural dermorphin were substituted by the corresponding α-methyl-β-azidoalanine or α-benzyl-β-azido(1-piperidinyl)alanine residues. The potency and selectivity of the new analogues were evaluated by a competitive receptor binding assay in rat brain using [3H]DAMGO (a μ ligand) and [3H]DELT (a δ ligand). The most active analogue in this series, Tyr-(R)-Ala-(R)-α-benzyl-β-azidoAla-Gly-Tyr-Pro-Ser-NH2 and its epimer were analysed by 1H and 13C NMR spectroscopy and restrained molecular dynamics simulations. The dominant conformation of the investigated peptides depended on the absolute configuration around Cα in the α-benzyl-β-azidoAla residue in position 3. The (R) configuration led to the formation of a type I β-turn, whilst switching to the (S) configuration gave rise to an inverse β-turn of type I′, followed by the formation of a very short β-sheet. The selectivity of Tyr-(R)-Ala-(R) and (S)-α-benzyl-β-azidoAla-Gly-Tyr-Pro-Ser-NH2 was shown to be very similar; nevertheless, the two analogues exhibited different conformational preferences.
New Article in ChemPhysChem
A strong case exists for the introduction of burst non-uniform sampling (NUS) in the direct dimension of NMR spectroscopy experiments. The resulting gaps in the NMR free induction decay can reduce the power demands of long experiments (by switching off broadband decoupling for example) and/or be used to introduce additional pulses (to refocus homonuclear coupling, for example). The final EXtended ACquisition Time (EXACT) spectra are accessed by algorithmic reconstruction of the missing data points and can provide higher resolution in the direct dimension than is achievable with existing non-NUS methods.
New Article in Journal of Biomolecular NMR
Alessandro Piai, Leonardo Gonnelli, Isabella C. Felli, Roberta Pierattelli, Krzysztof Kazimierczuk, Katarzyna Grudziąż, Wiktor Koźmiński, Anna Zawadzka-Kazimierczuk
Resonance assignment is a prerequisite for almost any NMR-based study of proteins. It can be very challenging in some cases, however, due to the nature of the protein under investigation. This is the case with intrinsically disordered proteins, for example, whose NMR spectra suffer from low chemical shifts dispersion and generally low resolution. For these systems, sequence specific assignment is highly time-consuming, so the prospect of using automatic strategies for their assignment is very attractive. In this article we present a new version of the automatic assignment program TSAR dedicated to intrinsically disordered proteins. In particular, we demonstrate how the automatic procedure can be improved by incorporating methods for amino acid recognition and information on chemical shifts in selected amino acids. The approach was tested in silico on 16 disordered proteins and experimentally on α-synuclein, with remarkably good results.
New article in Journal of Magnetic Resonance
Multidimensional NMR spectroscopy requires time-consuming sampling of indirect dimensions and so is usually used to study stable samples. However, dynamically changing compounds or their mixtures commonly occur in problems of natural science. Monitoring them requires the use multidimensional NMR in a time-resolved manner - in other words, a series of quick spectra must be acquired at different points in time. Among the many solutions that have been proposed to achieve this goal, time-resolved non-uniform sampling (TR-NUS) is one of the simplest. In a TR-NUS experiment, the signal is sampled using a shuffled random schedule and then divided into overlapping subsets. These subsets are then processed using one of the NUS reconstruction methods, for example compressed sensing (CS). The resulting stack of spectra forms a temporal “pseudo-dimension” that shows the changes caused by the process occurring in the sample. CS enables the use of small subsets of data, which minimizes the averaging of the effects studied. Yet, even within these limited timeframes, the sample undergoes certain changes. In this paper we discuss the effect of varying signal amplitude in a TR-NUS experiment. Our theoretical calculations show that the variations within the subsets lead to t1-noise, which is dependent on the rate of change of the signal amplitude. We verify these predictions experimentally. As a model case we choose a novel 2D TR-NOESY experiment in which mixing time is varied in parallel with shuffled NUS in the indirect dimension. The experiment, performed on a sample of strychnine, provides a near-continuous NOE build-up curve, whose shape closely reflects the t1-noise level. 2D TR-NOESY reduces the measurement time compared to the conventional approach and makes it possible to verify the theoretical predictions about signal variations during TR-NUS.
New article in Chemical Biology & Drug Design
Synthesis, biological activity and NMR-based structural studies of deltorphin I analogues modified in message domain with a new α,α-disubstituted glycines
Anika Lasota, Oliwia Frączak, Adriana Muchowska, Michal Nowakowski, Maciej Maciejczyk, Andrzej Ejchart and Aleksandra Olma
This paper describes new deltorphin I analogues in which phenylalanine residues were replaced by the corresponding (R) or (S)-α-benzyl-β-azidoalanine, α-benzyl-β-(1-pyrrolidinyl)alanine, α-benzyl-β-(1-piperidinyl)alanine and α-benzyl-β-(4-morpholinyl)-alanine residues. The potency and selectivity of the new analogues were evaluated by a competitive receptor binding assay in the rat brain using [3H]DAMGO (a μ ligand) and [3H]DELT (a δ ligand). The affinity of analogues containing (R) or (S)-α-benzyl-β-azidoalanine in position 3 to δ-receptors strongly depended on the chirality of the α,α-disubstituted residue. The conformational behavior of peptides modified with (R) or (S)-α-benzyl-β-(1-piperidinyl)Ala, which display the opposite selectivity, was analyzed by 1H and 13C NMR. The μ-selective Tyr-D-Ala-(R)-α-benzyl-β-(1-piperidinyl)Ala-Asp-Val-Val-Gly-NH2 lacks the helical conformation observed in the δ-selective Tyr-D-Ala-(S)-α-benzyl-β-(1-piperidinyl)Ala-Asp-Val-Val-Gly-NH2. Our results support the proposal that differences between δ- and μ-selective opioid peptides are attributable to the presence or absence of a spatial overlap between the N-terminal message domain and the C-terminal address domain.
New article in PLOS ONE
Sofia Unnerståle, Michal Nowakowski, Vera Baraznenok, Gun Stenberg, Jimmy Lindberg, Maxim Mayzel, Vladislav Orekhov, Tatiana Agback
Mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1) is a unique paracaspase protein whose protease activity mediates oncogenic NF-κB signalling in activated B cell-like diffuse large B cell lymphomas (ABC-DLBCLs). ABC-DLBCLs are aggressive lymphomas with high resistance to current chemotherapies. Low survival rate among patients emphasizes the urgent need for alternative treatment options. The characterization of the MALT1 will be an essential tool for developing new target-directed drugs against MALT1 dependent disorders. As the first step in the atomic-level NMR studies of the system, here we report, the 15N/13C/1H backbone assignment of the apo form of the MALT1 paracaspase region together with the third immunoglobulin-like (Ig3) domain, 44 kDa, by high resolution NMR. In addition, the non-uniform sampling (NUS) based targeted acquisition procedure is evaluated as a mean of decreasing acquisition and analysis time for larger proteins.
New article in Analyst
Monitoring polydispersity by NMR diffusometry with tailored norm regularisation and moving-frame processing
Nuclear magnetic resonance (NMR) is currently one of the main analytical techniques applied to numerous branches of chemistry. Furthermore, NMR has been proven useful to follow in-situ reactions occurring on a time scale of hours and days. For complicated mixtures, NMR experiments providing diffusion coefficients are particularly advantageous. However, the inverse Laplace transform (ILT) used to extract the distribution of diffusion coefficients from an NMR signal is known to be unstable and vulnerable to noise. Numerous regularisation techniques have been proposed to circumvent this problem. In our recent study, we proposed a method based on sparsity-enforcing l1-norm minimisation. This approach, which is referred to as ITAMeD, has been successful but limited to samples with a 'discrete' distribution of diffusion coefficients. In this paper, we propose a generalisation of ITAMeD using a tailored lp-norm (1 < p < 2 to process in particular signals arising from 'polydisperse' samples. The performance of our method was tested on simulations and experimental datasets of polyethylene oxides with varying polydispersity index. Finally, we have applied our new method to monitor diffusion coefficient and polydispersity changes of heparin undergoing enzymatic degradation in real-time.
New article in Polymer Chemistry
Controlling the stereoselectivity of rac-LA polymerization by chiral recognition induced formation of homochiral dimeric metal alkoxides
Paweł Horeglad, Martyna Cybularczyk, Anna Litwińska, Anna Maria Dąbrowska, Maciej Dranka,Grazyna Zukowska, Mateusz Urbańczyk and Michał Michalak
Using as an example dimeric dialkylgallium and dialkylindium alkoxide catalysts for the polymerization of rac-lactide (rac-LA), we have shown for the first time that formation of homochiral dimeric species [Me 2 MOR] 2 (M = Ga, In), induced by chiral recognition of monomeric Me 2 MOR units in the presence of Lewis base, leads to the increase of heteroselectivity of Ring Opening Polymerization (ROP) of rac-LA and therefore provides a new tool for controlling the stereoselectivity of polymerization of heterocyclic monomers. In order to explain the origin of heteroselectivity of [Me 2 Ga( μ -OCH(Me)CO 2 Me)] 2 /Lewis base system in ROP of rac-LA, the structure of (S,S)-[Me 2 Ga( μ -OCH(Me)CO 2 Me)] 2 ((S,S)-1) and rac-[Me 2 Ga( μ -OCH(Me)CO 2 Me)] 2 (1), in the absence and presence of tertiary amines and pyridines was investigated. The studies were further extended by the analysis of structure/activity studies of both (S,S)-[Me 2 In( μ -OCH(Me)CO 2 Me)] 2 ((S,S)-2) and rac-[Me 2 In( μ -OCH(Me)CO 2 Me)] 2 (2). Contrary to gallium complex 1, existing in solution as equimolar mixture of homo- and heterochiral diastereomers, in the case of 2 the excess of homochiral (R*,R*)-2 species was observed. For both Ga and In complexes the interaction of amine to metal center increased the tendency for the formation of homochiral species with the retention of dimeric structure in solution. Such a tendency was additionally demonstrated by the structure model dialkylgallium (3) and indium (4) complexes with monoanionic ligands possessing chiral center in α- position to alkoxide oxygen and pyridine functionality. The polymerization of rac-LA with gallium and indium catalysts (S,S)-1 and (S,S)-2 resulted in the formation of heterotactically enriched polylactide (PLA) (P r = 0.50 – 0.85) and (P r = 0.54 – 0.72), respectively. Heteroselectivity of investigated systems was in line with the excess of homochiral catalytic species. The higher activity of homochiral species activated by amines resulted in the positive non-linear effect between excess of homochiral (R*,R*)-1 or (R*,R*)-2 catalysts and heterotacticity of obtained PLA. The observed dependence of stereoselectivity of rac-LA polymerization on the excess of homochiral species was similar to the asymmetric amplification in enantioselective organic catalysis, however exceptional in polymerization processes.
New article in MRC
Fast acquisition of multidimensional NMR spectra of solids and mesophases using alternative sampling methods
New article in J. Phys. Chem. B
Lukasz Jaremko, Mariusz Jaremko, Michał Eligiusz Nowakowski, and Andrzej Ejchart
Nuclear magnetic relaxation provides a powerful method giving insight into molecular motions at the atomic resolution on a broad time scale. Dynamics of biological macromolecules has been widely exploited by measuring 15N and 13C relaxation data. Interpretation of these data relies almost exclusively on the use of the model-free approach (MFA) and its extended version (EMFA) which requires no particular physical model of motion and a small number of parameters. It is shown that EMFA is often unable to cope with three different time scales and fails to describe slow internal motions properly. In contrast to EMFA, genuine MFA with two time scales can reproduce internal motions slower than the overall tumbling. It is also shown that MFA and simplified EMFA are equivalent with respect to the values of the N-H bond length and chemical shift anisotropy. Therefore, the vast majority of 15N relaxation data for proteins can be satisfactorily interpreted solely with MFA.
New article in MRC
New article in Journal of Peptide Science
Analogues of deltorphin I containing conformationally restricted amino acids in position 2: structure and opioid activity
Anika Lasota, Oliwia Frączak,Anna Leśniak, Adriana Muchowska, Andrzej W. Lipkowski, Michał Nowakowski, Andrzej Ejchart and Aleksandra Olma
New analogues of deltorphin I (DT I, Tyr-d-Ala-Phe-Asp-Val-Val-Gly-NH2), with the d-Ala residue in position 2 replaced by α-methyl-β-azido(amino, 1-pyrrolidinyl, 1-piperidinyl or 4-morpholinyl)alanine, were synthesized by a combination of solid-phase and solution methods. All ten new analogues were tested for receptor affinity and selectivity to μ- and δ-opioid receptors. The affinity of analogues containing (R) or (S)-α-methyl-β-azidoalanine in position 2 to δ-receptors strongly depended on the chirality of the α,α-disubstituted residue. Peptide II, containing (S)-α-methyl-β-azidoalanine in position 2, displayed excellent δ-receptor selectivity with its δ-receptor affinity being only three times lower than that of DT I.
New article in Sensors
A group of signal reconstruction methods, referred to as compressed sensing (CS), has recently found a variety of applications in numerous branches of science and technology. However, the condition of the applicability of standard CS algorithms (e.g., orthogonal matching pursuit, OMP), i.e., the existence of the strictly sparse representation of a signal, is rarely met. Thus, dedicated algorithms for solving particular problems have to be developed. In this paper, we introduce a modification of OMP motivated by nuclear magnetic resonance (NMR) application of CS. The algorithm is based on the fact that the NMR spectrum consists of Lorentzian peaks and matches a single Lorentzian peak in each of its iterations. Thus, we propose the name Lorentzian peak matching pursuit (LPMP). We also consider certain modification of the algorithm by introducing the allowed positions of the Lorentzian peaks’ centers. Our results show that the LPMP algorithm outperforms other CS algorithms when applied to exponentially decaying signals.
New article in Analytical Chemistry
Nuclear magnetic resonance (NMR) spectroscopy is a versatile tool for chemical analysis. Besides the most straightforward application to study a stable sample containing a single compound, NMR has been also used for the analysis of mixtures. In particular, the analyzed mixtures can undergo changes caused by chemical reactions. The multidimensional NMR techniques are especially effective in a case of samples containing many components. Unfortunately, they are usually too lengthy to be applied in time-resolved experiments performed to study mentioned changes in a series of spectral “snapshots.” Recently, time-resolved nonuniform sampling (NUS) has been proposed as a straightforward solution to the problem. In this paper, we discuss the features of time-resolved NUS and give practical recommendations regarding the temporal resolution and use of the time pseudodimension to resolve the components. The theoretical considerations are exemplified by the application in challenging cases of fermenting samples of wheat flour and milk.
Cover in ChemPhysChem
The picture shows a spectral snapshot from a time-resolved two-dimensional NMR experiment with temperature sweep. The temperature is swept linearly in parallel to signal sampling. The new experiment proposed on p. 2217 by K. Kazimierczuk et al. allows precise, on-the-fly observation of various phase transitions. Dedicated processing with a compressed sensing algorithm provides a movie-like 2D spectrum that changes continuously with temperature.
Our EUROMAR 2014 contributions
- Talk: Thursday 15:00-15:20 HG F1 (PS320), Diffusion-ordered NMR with joint sparse sampling of time-gradient domain. Presenting: Mateusz Urbańczyk
- Poster: Monday 13:40-16:00 D-Floor A (ET443), Two-dimensional NMR spectroscopy with temperature sweep. Presenting: Rupashree Dass
- Poster: Wednesday 13:40-16:00 D-Floor A (ET444), Sparse sampling in non-frequency dimensions. Presenting: Krzysztof Kazimierczuk
Krzysztof Kazimierczuk was awarded with Wojciech Świętosławski Award from Polish Chemical Society.
New article in Angewandte Chemie
Accelerating Diffusion-Ordered NMR Spectroscopy by Joint Sparse Sampling of Diffusion and Time Dimensions
Diffusion-ordered multidimensional NMR spectroscopy is a valuable technique for the analysis of complex chemical mixtures. However, this method is very time-consuming because of the costly sampling of a multidimensional signal. Various sparse sampling techniques have been proposed to accelerate such measurements, but they have always been limited to frequency dimensions of NMR spectra. It is now revealed how sparse sampling can be extended to diffusion dimensions.
New article in Analyst
Criteria for sensitivity enhancement by compressed sensing: practical application to anisotropic NAD 2D-NMR spectroscopy
New article in ChemPhysChem
NMR like a movie: 2D nuclear magnetic resonance (NMR) spectroscopy is useful for studying temperature-dependent effects on molecular structure. However, the experimental time is usually long, because sampling is repeated at several temperatures. A novel solution to the problem is proposed, in which signal sampling is performed in parallel to the linear temperature sweep. Processing allows a movie, showing changes in a spectrum, to be created (see image).