NOESY transferred

Size restrictions are again similar to that encountered in protein NMR up to ca. 20-30 kDa complex size, line widths will usually be acceptable to allow detection of the XH resonances of the labeled or unlabeled component, including NOESY transfer steps. Beyond that, deuteration of the protein component becomes essential to reduce excessive line broadening. For the observation of the (15N-) labeled component (i.e., isotope editing), the use of TROSY techniques will further extend the size limit (Chapt. 10) however, this approach does not work for unlabeled components. 

Polarization transfer pathways for spectra (c) and (d) are depicted on a partial structure of 1 shown in the inset. Solid and dotted lines represent TOCSY and NOESY transfers, respectively. (Reprinted with adaptation with permission from ref. [39]. Copyright 1994 Academic... 

The proposed ID TOCSY-NOESY experiment is illustrated by the assignment of NOEs from anomeric protons H-lc and H-ld of the polysaccharide 1. Because the resonances of H-lc and H-ld overlapped, this assignment was not possible from a ID NOESY spectrum as shown in fig. 3(b). Although these protons differed in their chemical shifts, it was not possible to separate them by chemical-shift-selective filtration because of the very fast spin-spin relaxation of backbone protons (20-50 ms) in this polysaccharide. Instead, a ID TOCSY-NOESY experiment was performed in which the initial TOCSY transfer from an isolated resonance of H-2c was followed by a selective NOESY transfer from H-lc. The ID TOCSY-NOESY spectrum (fig. 3(c)) clearly separated NOE signals of the H-lc proton from those originating from the H-ld proton and established the linkage Ic —> 6a. 

Fig. 16. ID COSY-NOESY experiment on the polysaccharide 6 [77]. The structure of a terminal 3,6-dideoxy-4-C-(l-hydroxyethyl)-D-xylohexose is shown in the inset. The COSY transfer is depicted using the solid line, while a dotted line is used for the NOESY transfer, (a) H spectrum of 6 at 600 MHz and 50°C. (b) ID COSY-NOESY spectrum acquired using the sequence of fig. 13(c) with the initial transfer of magnetization from H-8 and the following parameters = 100 ms, to = 29 ms, Tr = 32 ms, A = 0.5 ms, N = 0, 1,...,64,...

In both ROESY mixing and TOCSY mixing, the transfer is from in-phase net magnetization on the spin-lock axis to in-phase net magnetization on the spin-lock axis for example, ly ->1 (TOCSY) or ly - - Iy (ROESY). This is analogous to the NOESY transfer, It -> — for small molecules and in contrast to the INEPT transfer 21 -> 21 1 , which is antiphase to antiphase. 

Tr-NOESY Transferred nuclear Overhauser effect spectroscopy... 

Figure 1.45 Coherence transfer pathways in 2D NMR experiments. (A) Pathways in homonuclear 2D correlation spectroscopy. The first 90° pulse excites singlequantum coherence of order p= . The second mixing pulse of angle /3 converts the coherence into detectable magnetization (p= —1). (Bra) Coherence transfer pathways in NOESY/2D exchange spectroscopy (B b) relayed COSY (B c) doublequantum spectroscopy (B d) 2D COSY with double-quantum filter (t = 0). The pathways shown in (B a,b, and d) involve a fixed mixing interval (t ). (Reprinted from G. Bodenhausen et al, J. Magn. Resonance, 58, 370, copyright 1984, Rights and Permission Department, Academic Press Inc., 6277 Sea Harbor Drive, Orlando, Florida 32887.)...

Both homonuclear and heteronuclear versions of relayed nOe experiments are known. The homonuclear relayed NOESY experiment involves both an incoherent transfer of magnetization between two spins H and H/ that are not coupled but close in space, and a coherent transfer of magnetization between two spins H(and H that are /-coupled together. The magnetization pathway may be depicted as... 

The NMR techniques discussed so far provide information about proton-proton interactions (e.g., COSY, NOESY, SECSY, 2D y-resolved), or they allow the correlation of protons with carbons or other hetero atoms (e.g., hetero COSY, COLOC, hetero /resolved). The resulting information is very useful for structure elucidation, but it does not reveal the carbon framework of the organic molecule directly. One interesting 2D NMR experiment, INADEQUATE (Incredible Natural Abundance Double Quantum Transfer Experiment), allows the entire carbon skeleton to be deduced directly via the measurement of C- C couplings. 

Oil and 0)2, and (b) 2D shift-correlation spectra, involving either coherent transfer of magnetization [e.g., COSY (Aue et al, 1976), hetero-COSY (Maudsley and Ernst, 1977), relayed COSY (Eich et al, 1982), TOCSY (Braunschweiler and Ernst, 1983), 2D multiple-quantum spectra (Braun-schweiler et al, 1983), etc.] or incoherent transfer of magnedzation (Kumar et al, 1980 Machura and Ernst, 1980 Bothner-By et al, 1984) [e.g., 2D crossrelaxation experiments, such as NOESY, ROESY, 2D chemical-exchange spectroscopy (EXSY) (Jeener et al, 1979 Meier and Ernst, 1979), and 2D spin-diffusion spectroscopy (Caravatti et al, 1985) ]. 

A 3D spectrum of a three-spin subsystem in which all the nuclei are coupled to one another, such as C(H/i)(Hb)-C(Hc), will lead to 27 peaks, comprising six cross-peaks, 12 cross-diagonal peaks (six at o) — o>2 and the other six at 0)2 = (O3), six back-transfer peaks, and three diagonal peaks. However, in the case of a linear three-spin network (e.g., CH -CHg-CHc), the number of peaks will depend on whether two equal (e.g., COSY-COSY or NOESY-NOESY) or unequal (e.g., COSY-NOESY) mixing processes are... 

In the case of a COSY-NOESY spectrum having an unequal mixing, let us consider two nuclei (say, A and B) that are spatially close but belong to different coupling networks, and nuclei B and C, which have scalar coupling with each other but are spatially distant. The only transfers allowed in this situation are ... 

The pulse sequences for HMQC-COSY and HMQC-NOESY experiments are presented in Fig. 6.10. The 3D HMQC-COSY spectrum of a N labeled tripeptide is shown in Fig. 6.11. Since the coherence transfer involved in this experiment is N([Pg.362]

Two-dimensional relayed NOESY experiments (Wagner, 1984 Kessler et al., 1988) give cross-peaks that could be a superposition of nOe s resulting from different relay nuclei. This complicates the extracting of crossrelaxation rates. The 1D NOESY experiment (Kessler et al., 1989a), however, allows the path of the magnetization transfer to be clarified. 

Fig. 9.1 The internuclear transfer of magnetization via NOE cross-relaxation in an isolated spin-pair. (A) Build-up curves for the cross-peak intensity in a 2D NOESY experiment for various internuclear distances r. The dashed line indicates a typical mixing time tm = 300rns used for drug-like molecules.

Moseley, H. N. B., Curto, E. V., Krishna, N. R. Complete Relaxation and Conformational Exchange Matrix (CORCEMA) analysis of NOESY spectra of interacting systems two-dimensional transferred NOESY. J. Magn. Reson. Sen B 1995, 108, 243-261. 

---