Statut actuel de NUS et APSY

Transcription

1 Statut actuel de NUS et APSY Martial PIOTTO Bruker BioSpin, France 30 ème Réunion Utilisateurs Novembre 2016 Innovation with Integrity

2 General features of NUS and APSY Techniques that allow to collect n-dimensional data without the need for a complete sampling of the points in the indirect dimension(s) Non-Uniform Sampling: Technique used for n 2 (2D/3D/4D) data Small molecules Biological applications (Proteins ) APSY: Technique used for n 3 (3D 7D...) data Biological applications (Proteins )

3 Different sampling schemes (3D experiment / Two indirect dimensions) Traditional sampling NUS sampling Radial Sampling (APSY) From: Kazimierczuk, K., V. Orekhov (2015). Nonuniform sampling: post-fourier era of NMR data collection and processing. MRC 53,

4 Non Uniform Sampling (NUS)

5 Acquisition of NUS data standard since TopSpin 3.0 Non-uniform sampling scheme (3D acquisition with two indirect dimensions) t 2 t 2 50 complex points t 1 t 1

6 Data Processing Implementation in Topspin 3.5 pl6 Multidimensional decomposition MDD-NMR Maximum Entropy (MaxEnt) Rowland Toolkit Forward Maximum Entropy Azzara (CCPN) Multidimensional Fourier Transformation MFT Compressed Sensing CS Orekhov et al. Hoch et al. Wagner et al. Laue et al. Kozminski et al. Marion Orekhov et al. Kazimierczuk Nietlispach et al.

7 Data Analysis Implementation in Topspin 3.5 pl6 Compressed Sensing (Two different algorithms) CS IST CS IRLS With/Without Virtual Echo 2D/3D Multidimensional decomposition MDD-NMR (Recursive and non-recursive) 3D/4D Parallel processing for: Linux Mac Windows

8 Full integration of NUS techniques on the spectrometer Acquisition «eda»

9 Full integration of NUS techniques on the spectrometer Processing «edp» Available options: MDD recursive MDD (R-MDD) CS IST (Iterative Soft Thresholding) CS IRLS (Iterative Reweighted Least Squares) All published parameters available through the command line

10 NUS Results Increased resolution in f1 for HSQC Ubiquitine 2 mm Spectral enhancement with Non-Uniform-Sampling Ubiquitine 2 mm 2D 1H-15N HSQC 10 min acquisition time CS processing Traditional sampling 256 points NUS sampling 1024 points NUS 25%

11 NUS Results Increased resolution for 3D 15N NOESY-HSQC Ubiquitine 2 mm Regular sampling 50x100 complex points in 14h37 ppm NUS sampling 20% 50x256 complex points in 7h23 ppm ppm ppm

12 NUS Results Increased resolution for 3D 15N NOESY-HSQC Ubiquitine 2 mm Regular sampling 50x100 complex points in 14h37 ppm NUS sampling 20% 50x256 complex points in 7h23 ppm ppm ppm

13 NUS Results Increased resolution for 3D 15N NOESY-HSQC Ubiquitine 2 mm NUS sampling 20% 50x256 complex points in 7h23 Regular sampling 50x100 complex points in 14h ppm

14 Practical considerations when acquiring and processing NUS 2D experiments For 2D experiments, CS is the method of choice The number of complex data points acquired in the indirect dimensions should be similar to the number of expected cross-peaks Example: 2D 1H/15N HSQC experiment on ubiquitine: About 100 cross-peaks expected 128 complex points acquired For 2D experiments, NUS is particularly powerful to record 2D HSQC and HMBC spectra Typically gain of factor of 2 in acquisition time (50 % NUS) Higher resolution in the same amount of time Use relaxation matched sampling (according to estimated T2) Phasing a 2D spectrum processed with NUS Need to regenerate the imaginary part in f2 with a Hilbert transform (xht2)

15 Practical considerations when acquiring and processing NUS nd experiment (n>2) Compressed Sensing (CS) processing can be computer demanding for 3D spectra (3D NOESY-HSQC experiments) MDD processing is less computer demanding (4D data set) When possible, limit the processing to the region of interest using STSR and STSI (i.e. for proteins limit the processing to the NH) Phasing a nd spectrum processed with NUS Need to regenerate the imaginary parts with a Hilbert transform (xht2 ; xht1 )

16 Data Analysis NUS licenses with Topspin 3.5 pl6 NO LICENSES FOR: Compressed Sensing CS IST for 2D experiments without virtual echo SINGLE LICENCE REQUIRED FOR: Compressed Sensing CS IST (>2D experiments) CS IRLS With/Without Virtual Echo Multidimensional decomposition MDD-NMR (Recursive and non-recursive)

17 APSY Automated Projection SpectroscopY Hiller S, Fiorito F, Wüthrich K, Wider G. Automated projection spectroscopy (APSY). PNAS 2005; 102(31): 10876

18 Principles of Automated Projection Spectroscopy (APSY) - Technique to accelerate the acquisition and analysis of multidimensional data (n 3) a=0 - Procedure: Record 2D projections of high-dimensional experiments Automatic peak picking of the projections Geometric analysis of the peak list Generation of a n-dimensional list of crosspeaks (3D 3 frequencies) a=90 a E. Kupce and R. Freeman, J. Am. Chem. Soc., 126, 6429, Hiller S, and Wider G., Topics in Current Chemistry; 2012

19 APSY: General Procedure as implemented in Topspin3.5pl6 - Select experiment - Generate a list of projection angles - Acquire corresponding projections as 2D planes - Process 2D planes - Analyze 2D planes - Generate peak list and report Hiller S, and Wider G., Topics in Current Chemistry; 2012, p

20 APSY: General Procedure as implemented in Topspin3.5pl6 Uses the acquisition mode (Fn-Type) PROJECTION SPECTROSCOPY Compatibility of Bruker library pulse programs with APSY mode of acquisition No need for specific APSY PP anymore Standard Pulse Program from the Bruker library can be used Full integration in TopSpin (like NUS)

21 APSY in Topspin3.5 pl6 Standard PP

22 APSY in Topspin3.5 pl6 apsy command

23 APSY in Topspin3.5 pl6 apsy toolbar Angles: Create list of projection angles GAPRO: Parameters used for peak identification (S/N ) Setup: Create all the 2D experiments Run: Runs all the different 2D experiments Stop: Stop Re-Process: Allows to automatically reprocess the entire series of spectra (Phase ) Re-Evaluate: Re-evalute the newly processed data Results: Peak list Help: Help

24 APSY in Topspin 3.5 pl6 Results

25 APSY: General Features APSY can be used to record 3D, 4D, 5D and 6D experiments Application Intrinsically disordered proteins (IDP) NUS processing limited to 2D, 3D and 4D NUS processing times can become important for high dimensionality spectra

26 APSY: Results 3D + 5D + 5D experiments High precision peak lists HNCA 3D HACACONH 5D CBCACONH 5D N CA NH HA CA CO N NH CA CB CO N NH Dutta, S.; Serrano, P.; Proudfoot, A.; Geralt, M.; Pedrini, B.; Herrmann, T.; Wüthrich, K. Journal of Biomolecular NMR 2015, 61,

27 APSY experiments 6D sequential H i -N i -CO i-1 -CA i-1 -N i-1 -H i-1 d H N i a C a i-1 N i e N i-1 c C i-1 b H N i-1 O

28 APSY experiments Ubiquitin 2mM Key Features: High Precision: Peak lists of high precision from an optimum number of projections. H i-1 N i-1 CO CA N i H i (N i-1 -N i ) ppm ppm ppm ppm ppm ppm Hz Sequential assignment of [ 13 C, 15 N]-ubiquitin using the peak list from a 6D-APSY-HNCOCANH experiment. Dutta, S.; Serrano, P.; Proudfoot, A.; Geralt, M.; Pedrini, B.; Herrmann, T.; Wüthrich, K. Journal of Biomolecular NMR 2015, 61,

29 Acknowledgements Detlef Moskau Wolfgang Bermel W. Mausshardt E. Schweitzer V. Orekhov V. Jaravine S. Hiller G. Wider

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