Q-HSQC

Heikkinen et al. presented an application of the Q-HSQC experiment with uniform polarization transfer over a range of cn couplings in quantification. This study with a mixture of lignin model compounds demonstrated comparative... 

Figure 11.20 shows three F2 slices of the HSQC spectrum, at the F shifts of carbons j, q, and s. For q and s we see the classic doublet-triplet pattern, but for j there is a broad,... 

Figure 12.51 shows the 2D XH, 15N HSQC spectrum of U-15N Rhodobacter capsu-latus Ferrocytochrome q in 90% H2O. The F planes of the corresponding 3D TOCSY-HSQC spectrum are indicated on the left side of the 2D spectrum. Plane 15 (lower dotted line) corresponds to a 15N chemical shift of 131.59 ppm, very close to the 15N shift of the backbone nitrogen of Ala-66 (131.66). Plane 16 (upper dotted line) corresponds to... 

Figure 12.12a gives a good illustration of the need for going to a third dimension to facilitate the interpretation of a crowded 2D spectrum. The NOESY spectrum of a uniformly 15N-enriched protein, staphylococcal nuclease, has so many cross peaks that interpretation is virtually impossible. However, it is possible to use, 5N chemical shifts to edit this spectrum, as indicated in Fig. 12.121) and c in a three-dimensional experiment. With the 15N enrichment, NOESY can be combined with a heteronuclear correlation experiment, in this case HMQC, but HSQC could also be used. A 3D pulse sequence can be obtained from two separate 2D experiments by deleting the detection period of one experiment and the preparation period of the other to obtain two evolution periods (q and t2) and one detection period (f3). In principle, the two 2D components can be placed in either order. For the NOESY-HMQC experiment, either order works well, but in some instances coherence transfer proceeds more efficiendy with a particular arrangement of the component experiments. We look first at the NOESY-HMQC sequence, for which a pulse sequence is given in Fig. 12.13. The three types of spins are designated I and S (as usual), both of which are H in the current example, and T, which is 15N in this case.

Similar to the HSQC experiment, multiple quantum coherences can be used to correlate protons with Q-coupled heteronuclei. The information content of the Heteronuclear Multiple Quantum Correlation (HMQC) experiment (56) is equivalent to the HSQC, but the sensitivity can be improved in certain cases. Additionally, by proper tuning of delays and phase cycling, it can be transformed into the heteronuclear multiple bond correlation experiment (57-59), which results in correlations between J- and J-coupled nuclei. 

Figure 14. A comparison of three HETCOR spectra of naturally abundant L tyrosine-HCI recorded under as far as possible equivalent conditions using (a) the FSLG-decoupled CP (Figure 11), (b) the MAS / HMQC (Figure 12), and (c) the REPT-HSQC (Figure 13) experiments. Above each spectrum, the 13C skyline projection as well as the Fourier transformation of the first slice (q = 0) of the 2D data set is shown, while below each spectrum the 11 i skyline projections are shown. The spectra were recorded at a H Larmor frequency of 500 MHz the improvement in resolution at a Larmor frequency of 700 MHz for the REPT-HSQC experiment is shown as a dashed line. (Reproduced with permission from ref 125. Copyright 2001 Academic Press.)...

Heteronuclear coupling constants (1,b7c,h) are most commonly measured from heteronuclear 2D experiments. The 3/c H couplings can be easily extracted from /-resolved spectra as well as from f or F2 proton coupled HSQC spectra. The undesired evolution of "/CH during q can be eliminated with use of an appropriate bilinear rotation decoupling (BIRD) pulse, such as BIRDd,x in. /-resolved spectroscopy35 and 111RD in Fi-coupled HSQC.36 Spin-state selective excitation techniques, S3E and S3CT37 38 (spin-state-selective coherence transfer), can also be used for the measurement of... 

Fig. 7. Selected region from the H-coupled H- N HSQC spectrum of 1 mM TwCsp in isotropic solution (A) and in a solution containing 5 wt.-% DMPC/DHPC bicelles (q = 3.0, CTAB-doped) measured at 336 K and a pressure of 150 MPa. The effective H- N coupling constant giving rise to the doublet splitting in indirect spectral dimension (5i) is given together with the assignment of the signals. (After ref 71.)...

The 3D structure of sensory rhodopsin II (pSRII) solubilized in detergent (DHPC) or biceUe q = 0.3) was determined by solution NMR [66]. The overall quaHty of the structure ensemble is good (backbone r.m.s. deviation of 0.48 A) and agrees well with previously determined X-ray structures. The local and global dynamics of a chemokine receptor CXCRl (350-residue GPCR) are characterized using a combination of solution NMR and soHd-state NMR experiments [67]. In isotropic bicelles ( = 0.1), only 13% of the expected number of backbone amide resonances is observed in HSQC NMR spectra of uniformly N-labelled sam-... 

An exhaustive set of residue type selective triple resonance experiments have be proposed recently. They all include MUSIC multiplet filter in combination with a multi-step coherence transfer to observe sequential (i -I- 1)-HSQC or intraresidue and sequential (i,i -I- 1)-HSQC cross-peaks separately for different residue types. Four additional experiments were added to the repertoire Gin-specific Q-(i -I- l)-HSQC/Q-(i,i -I- 1)-HSQC that apply MUSIC selection to... 

Figure 8 Basic pulse schemes to obtain F2-heterocoupled two-dimensional HSQC spectra (A) CLIP-HSQC, (B) perfect-CLIP-HSQC, and (Q PIP-HSQC experiments. Narrow and broad pulses represent 90° and 180° pulses, respectively, with phase x, unless specified explicitly. The interpulse delay A is set to 1/(2 J(CH)) and a basic two-step phase cycling is executed with i=x,-x and receiver (fi,=x—x. Gradients for coherence selection using the echo-antiecho protocol are represented by G1 and G2 and 8 stands for the duration and the gradient and its recovery delay. A purge gradient G3 is placed for zz-filtering whereas the final and optional 90°O C) stands for the so-called CLIP pulse to remove heteronuclear AP contributions. F2-heterodecoupled versions of all three HSQC schemes should be obtained by applying broadband heterodecoupling during the acquisition period. In such cases, the CLIP pulse In (A) and (B) is not required.

In the 2D-NMR spectra, carbon resonances are labeled a-q and regions of the proton spectram are labeled 1-4, with assignments alongside the labels. Crosspeaks are identified by letter and numerical subscript. Carbon resonance 1 does not contain HMBC or HSQC-TOCSY crosspeaks to CHooe/eoo or CHooo, thus this signal must be attributed to ay carbons in EOEOE pentads. Based on... 

---