Resonance assignments

H and UCNMR spectral data for l//-azepin-3(2//)-ones have been discussed in detail and resonances assigned.86 Similar data are available for 3-hydroxyazepin-2-ones and their 2-and 3-0-alkyl ethers.48 51... 

The partial backbone assignment of the catalytic domain of protein kinase A (PKA) using a combination of NMR techniques was reported in 2004 [51]. This represented the first reported NMR assignment of any protein kinase catalytic domain. Backbone resonance assignment of the 42kDa protein was achieved using a combination of triple-labelled ( H, N)... 

Fig. 2 Chemical shift perturbation and chemical shift mapping, (a) Portions of the [15N, 1H]-HSQC spectra of Bcf-xL recorded in absence (black) and in presence of each of the four molecules (in colors). Resonance assignments for amino acid residues that exhibit large shifts are reported, (b) Structure of Bc1-Xl in complex with the BH3 peptide from Bak (PDB code 1BXL) showing the chemical shift changes in Bcl-xL upon ligand binding (blue, large shits yellow, no shifts the Bak peptide is reported in cyan). Adapted from [48]...

On the other hand, resonance assignments for CP of threonine and serine, and C and Cy of hydroxy proline, were difficult to make, because of their proximity to carbohydrate carbon resonances. In most cases then, the resonances were assigned on the basis of the effects of pH on the chemical shifts of those resonances. It was shown that the chemical shifts for the carbohydrate carbon resonances were virtually unaffected (AS 0.4 p.p.m.) when going from the cationic state (pH 2) to the anionic state (pH 11) of the amino acid residues. The chemical shifts of C and CP of the amino acid residues, however, shifted considerably (up to 3.1 and 6.6 p.p.m. for C" and CP, respectively see Table VI). 

Now that the general considerations concerning the 13C-n.m.r.-spec-tral data for O-glycosylated model compounds have been discussed, attention may be focused on some specific cases. Table IV gives the 13C-n.m.r.-spectral data for L-serine that has been glycosylated with various carbohydrates, such as a- and / -d-G1c, a- and /9-D-Gal, a- and / -d-Xyl, a-D-Man, and a-D-GalNAc. As mentioned earlier, the specific resonance-assignments to amino acid carbon atoms were based on chemical... 

C-N.m.r. data for only a limited number of model compounds of this nature are currently available (see later). The 13C-n.m.r. data for the a-D-Man — Ser linkages, given in Tables IV and V, have already been discussed. Their application to 13C-spectral analysis has been shown in Figs. 1 and 4, and in the discussion of the glycoprotein glucoamylase. They allowed specific, resonance assignments to be made for the respective spectra. 

Zhang O, Kay LE, Olivier JP, Forman-Kay JD. Backbone 11 and 15N resonance assignments of the N-terminal SFI3 domain of drk in folded and unfolded states using enhanced sensitivity pulsed field gradient NMR techniques. J Biol NMR 1994 4 845-858. 

Table 3. Proton nuclear magnetic resonance assignments for spectrum of sodium valproate...

Fig. 1. Typical values for the scalar couplings in perdeuterated proteins, which can be used for the coherence transfer through the spin system (a). Strategies for obtaining resonance assignment in 15N/13C/2H labelled proteins discussed in this chapter (b). The boxes, circles, and triangles indicate correlations established in the corresponding pulse scheme.

The effect of deuteration, for the needs of sequence-specific resonance assignments using triple-resonance experiments, was first demonstrated by Bax and co-workers on the 19.7 kDa protein calcineurin B in 1993.56 By replacing the H spin with deuterium, the transverse relaxation time of 13C spin is increased by nearly an order of magnitude due to the 6.5 times smaller gyromagnetic ratio of 2H in comparison to in.52,56 Not surprisingly, deuteration was utilized for the aid of structure determination of several... 

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