Thesis Title: Subtitle - NMR Spectroscopy Research Group

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46 Chapter 2. Possum: paramagnetically orchestrated spectral solver of unassigned methyls. d Fraction of correct assignments for the paramagnetic cz- 186/ /Yb 3+ and cz- 186/ /Dy 3+ complexes, as indicated. The number in the denominator is the number of methyl groups for which cross-peaks were observed in the presence of Yb 3+ or Dy 3+ . e Fraction of correct assignments for the diamagnetic cz- 186/ /La 3+ complex. The number in the denominator is the number of methyl groups for which cross-peaks were observed in at least one of the paramagnetic complexes. 2.4.6 PCS and flexibility Structural differences between the crystal structure of 186 determined under cryogenic conditions (Hamdan et al., 2002b) and the solution structure of the cz- 186/ complex become apparent as differences between measured and predicted PCS. In a few cases, the structural differences interfere with the resonance assignment. Figure 2.6a illustrates the situation for the cz- 186/ /Dy 3+ complex, where the measured PCS of Leu161 are smaller than predicted and would more closely match the values predicted for Leu131. This can be explained by a small displacement of the peptide segment comprising residues 151-161 that decreases the PCS of both methyls of Leu161. Smaller PCS than expected were also observed for the backbone amides of this segment. 45 The correct assignment would be obtained by focusing on the difference in 13 C PCS between both methyl groups rather than their magnitude (Figure 2.6a) or by using the data of the cz- 186/ /Yb 3+ complex which are less strongly distance dependent in the 11 Å distance range (Figure 2.6b).
2.4 Results. 47 Figure 2.6 Residues showing deviations between predicted and experimental PCS. Comparison of calculated and experimental PCS of Leu131, Leu95, Leu161, Leu11 and Ile154 in the cz- 186/ /Dy 3+ complex. The data are plotted in the sequence C 1 /H 1 /C 2 /H 2 and C 2 /H 2 /C 1 /H 1 / for the Leu residues and Ile154, respectively. (b) Same as (a), but for the cz- 186/ /Yb 3+ complex. (c) Predicted 13 C PCS of the prochiral methyl groups of Val82 in cz- 186/ /Dy 3+ versus sidechain dihedral angle. The values predicted from the crystal structure of 186 40 are marked. (d) Same as (c), but for the CH3 groups of Leu95. In the cases of Val82 and Leu95 in the Dy 3+ complex, the comparison of experimental and predicted 13 C-PCS data yields the wrong stereospecific assignment. The 1 and 2 angles of these residues are –47º and 172º, respectively, in the crystal structure (Hamdan et al., 2002b). Adjusting these angles to –60º and 180º, respectively, inverts the relative size of the 13 C PCS predicted for the two methyl groups, leads to much better agreement between predicted and experimental PCS, and results in the correct stereospecific assignments (Figure 2.6c and d). This observation is most simply explained by a small difference between the crystal and solution structures. Note that the
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Computational study of proteins wit
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iii Schmitz C, Stanton-Cook MJ, Su
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v Acknowledgements I would like to
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vii Keywords paramagnetic nmr, pseu
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ix 2.3.3 Manual resonance assignmen
Page 11 and 12: xi List of Figures Figure 1.1 NMR e
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3.11 Supporting information. 97 Zwe
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102 Chapter 4. Protein Structure De
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Chapter 5 Conclusion and perspectiv
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5.1 The use of PCS for structure de
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5.1 The use of PCS for structure de
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5.3 The use of PCS for protein dock
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