Resonance assignment of the apoprotein was carried out by using available 1H and 15N chemical shift data [66], [67], with the aid of 2D TOCSY and NOESY, and 3D 15N-edited NOESY spectra.
Glucosylation positions of the kaempferol and quercetin reaction products were assigned based on the hypsochromic shift data [ 50], comparison with published data [ 31, 47, 51] and when available, using authentic reference compounds.
To check if positioning of Ub into the catalytic site could be consistent with the measured chemical shift data, the three runs were repeated with one additional distance restraint (10 Å) between the Josephin side chain sulphur of C14 and the C-terminal carbon of site 2.
Comparison of QM/MM-calculated NMR chemical shifts with experimental shift data for a selective subset of backbone 15N provides an additional evaluation criteria.
The combination of NMR relaxation experiments with chemical shift data and the linkage to structure/function reveals a fundamentally different activation mechanism.
The spectrum was assigned, and the sites of N- and O-sulfation and the conformation of each uronic acid residue were established, with chemical shift data obtained from BASHD-TOCSY spectra, while the sequence of the monosaccharide residues in the octasaccharide was determined from inter-residue NOEs in BASHD-NOESY spectra.
The proposed mechanism is consistent with NMR chemical shift data at increasing temperature and recent experimental observations pointing to a pivotal role of secondary structure elements in directing the folding process toward the native state.
The chemical shift data are available at the Biological Magnetic Resonance Data Bank with accession number 16616.
> -wrap-foot>The sites of N-sulfation and O-sulfation were then established using reference chemical shift data for heparin monosaccharides [ 20], with the result that the octasaccharide was determined to have the sequence ∆UA 2S -[(1 → 4)-GlcNS(6S)-(1 → 4)-IdoA(2S)]3-(1 → 4)-GlcNS(6S) (Fig. 4).
Tested on non-mutant, full-length amyloid structures with known chemical shift data, AmyloidMutants offers roughly 2-fold improvement in prediction accuracy over existing tools.
