DQF-COSY Double-quantum filtering

NMR spectroscopy was carried out using a Varian Unity 300MHz spectrometer. Peptides were dissolved in 500 pL of 90% H,O/10% D20 (or 100% D20) giving a sample concentration of 1-2 mM and the pH adjusted to 5.5. H DQF-COSY (double quantum filtered two-dimensional correlated spectroscopy), ROESY, and TOCSY spectra were collected at 25 °C and processed as described.1 6-281... 

DQF-COSY (double quantum filtered correlation spectroscopy) 157,158... 

Lin et al. have reported (p and angles for the sLc tetrasaccharide, determined with the aid of ROESY NMR measurements and Molecular Mechanics 2 (MM2) calculations. as NeuAc. Gal 163°, —61°, Gal, GlcNAc 48°, 15° andFuc, GlcNAc 22°, 30°. In a subsequent study from the same laboratory, Ichikawa etal. used DQF-COSY (double quantum-filtered correlation spectroscopy) and TOCSY (total correlation spectroscopy) to derive complete H and C... 

DQF-COSY Double-quantum filtered correlation spectroscopy 5.6.2... 

List of Abbreviations NMR, nuclear magnetic resonance O.D.260> optical density at 260 nm at 25 °C NOESY, nuclear Overhauser effect spectroscopy DQF-COSY, double quantum filtered correlation spectroscopy KP, potassium phosphate Tm, UV-melting point ER, endoplasmic reticulum PAGE, polyacrylamide gel electrophoresis TSP, trisilylpropionic acid sodium salt fid, free induction decay ffh, fifty-four-homologue j57iAf, methionine-rich domain of fifty-four-homologue. 

In principle, 2D NMR structural determination of proteins in solution involves the linking of information derived from a combination of three complementary experiments - TOCSY (Total Correlated Spectroscopy), DQF-COSY (Double-Quantum Filtered Correlated Spectroscopy) and NOESY (Nuclear Overhauser Effect Spectroscopy). The TOCSY (58) spectra are used to identify spin systems of the amino acids in the protein. COSY (59,60) spectra yield complementary information to the TOCSY, which can be used to obtain the direct scalar connectives. The NOESY (61) spectra yield information on the through space relationship of protons in the protein. It should be noted that because of the vast abundance of hydrogen atoms in a protein molecule proton NMR is the preferred technique to determine the solution structure of a protein. 

A suit of 2D-NMR techniques that include chemical-shift correlation spectroscopy (COSY), double-quantum filtered correlation spectroscopy (DQF-COSY), total correlation spectroscopy (TOCSY), and heteronuclear multiple-quantum coherence spectroscopy (HMQC) have been used for... 

Suppression of the tme diagonal peaks by double-quantum filtering (DQF-COSY) may resolve such problems. Finally, quantitative measurements of the magnitude of the coupling constants is possible using the Z-COSY modification, These experiments ate restricted to systems of abundant spins such as H, and which have reasonably narrow linewidths. 

Figure 5.37 (a) Conventional phase-sensitive COSY spectrum of basic pancreatic trypsin inhibitor, (b) Double-quantum filtered (DQF) phase-sensitive COSY spectrum of the same trypsin inhibitor, in which singlet resonances and solvent signal are largely suppressed. Notice how clean the spectrum is, especially in the region near the diagonal line. (Reprinted from Biochem. Biophys. Res. Comm. 117, M. Ranee, et al., 479, copyright (1983) with permission from Academic Press, Inc.)... 

The DQF (double-quantum filtered)-COSY spectrum of an isoprenyl coumarin along with H-NMR data are shown. Determine the H/ H homonuclear interactions in the DQF-COSY spectrum. 

The data from H NMR studies of 63, which included double quantum filtered phase sensitive correlated spectroscopy (DQF-COSY) and rotating frame nuclear Overhauser effect spectroscopy (ROESY) experiments (Figure 12), are collected in Table 17. 

FIGURE 5.10 Pulse sequence for Double Quantum Filtered H—Tf COSY (DQF -COSY). 0 s are 90° pulses and 8 is a fixed delay of the order of a few microseconds. 

The DQF H— H COSY spectrum of ipsenol can be found at the bottom of Figure 5.9. Note that the spectrum seems cleaner, especially along the diagonal, making the task of interpretation significantly easier. Because of the greatly improved appearance of DQF-COSY, all COSYs in this book are double quantum filtered. 

The MQC intermediate state in coherence ( INEPT ) transfer can also be used to clean up the spectrum. In this case, we can apply a double-quantum filter (using either gradients or a phase cycle) to kill all coherences at the intermediate step that are not DQC. We will see the usefulness of this technique in the DQF (double-quantum filtered) COSY experiment (Chapter 10). As with the spoiler gradient applied to the 2IZSZ intermediate state, a doublequantum filter destroys any unwanted magnetization, leaving only DQC that can then be carried on to observable antiphase magnetization in the second step of INEPT transfer. 

A simple variant of the COSY experiment is COSY-35 (sometimes called COSY-45), in which the second 90° pulse is reduced from a 90° pulse to a 35° or 45° pulse (Fig. B.3). The result is that the fine structure of crosspeaks is simplified, with half the number of peaks within the crosspeak. This makes it much easier to sort out the coupling patterns in both dimensions and to measure couplings (active and passive) from the crosspeak fine structure. A more important variant of the COSY experiment is the DQF (double-quantum filtered)— COSY (Fig. B.4), which adds a short delay and a third 90° pulse. The INEPT transfer is divided into two steps antiphase I spin SQC to I,S DQC, and I,S DQC to antiphase S spin SQC. The filter enforces the DQC state during the short delay between the second and third pulses either by phase cycling or with gradients. DQF-COSY spectra have better phase characteristics and weaker diagonal peaks than a simple COSY, so this has become the standard COSY experiment. 

Double quantum filters can be used as a component of many different 2D, 3D, and 4D experiments. In some applications, COSY benefits from DQF, as we discuss in the next section. 

The structural connectivity derived from examination of the 111, 13C/DEPT, DQF-COSY, HMQC, and HMBC data (DEPT = distortionless enhancement by polarization transfer DQF = double quantum filtering COSY = correlation spectroscopy HMQC = heteronuclear multiple quantum correlation HMBC = heteronuclear multiple bond correlation) resulted in global reevaluation of sclerophytin B structure and demonstrated that this compound and the related alcohol are not composed of two ether bridges as in the originally formulated structure 37, but share the structural features depicted as 38 <20000L1879>. Comparison of 13C and 111 NMR data of Norte s... 

H, C, and NMR chemical shifts of 10 substituted pyrazolo[l,5- ]pyrimidines 40 were assigned based on double quantum filtering (DQF) H, H correlation spectroscopy (COSY), pulsed field gradient (PFG) H, C... 

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