Thesis Title: Subtitle - NMR Spectroscopy Research Group

More documents

Recommendations

Info

12 Chapter 1. Introduction. 1.3 Computational study of paramagnetic proteins 1.3.1 The assignment problem Assigning the resonances of <strong>NMR</strong> spectra is a necessary step towards applying <strong>NMR</strong> restraints for protein computation. Although several software packages are capable of predicting chemical shifts, they require high-resolution 3D structures and lack accuracy especially for the unstructured parts of a protein (Shen et al., 2007). Their algorithms are not based on pure calculation from the 3D structure, but rather use statistical information extracted from the pdb data bank and from deposited chemical shifts. Pseudocontact shifts can be accurately predicted using equation (1.1). Consequently, it is possible to compare measured and calculated PCSs. The root mean square deviation between the calculated and the measured PCSs provides a score to minimize in order to yield the best possible assignment. This strategy has been applied to simultaneously assign measured PCSs and optimize the Δχ-tensor by a software package named ―Platypus‖ (Pintacuda et al., 2004). In this work, the protein was selectively labeled by residue type in order to simplify spectra. This led to the measurement of unassigned PCSs by unambiguous identification of the connectivity between a diamagnetic cross peak and its paramagnetic partner shifted along the diagonal in a 2D 15 N-HSQC spectrum. The diagonal shift is explained by the fact that in first approximation, hydrogen and nitrogen of an NH group have similar PCS values. This is due to the short distance between N and H atoms (approximately 1 Å) compared to the large distance (at least 10 Å) separating the NH bond from the lanthanide inducing the observed PCS. As a result, the polar coordinates within the Δχ- tensor frame are similar for the N and H spins and hence, both spins experience similar pseudocontact shifts. The second step of the protocol consists of combining a grid search over the Δχ-tensor parameters with an optimal assignment algorithm called the Hungarian method (Kuhn, 1955): The grid search covers a large ensemble of possible combinations for the Δχ-tensor parameters. At each node of the grid search, it becomes possible to use the Hungarian method to obtain the optimal assignment in a polynomial time. A score can be calculated over each node to reflect the quality of the assignment, and compared to other nodes to extract the best assignment along with the best set of Δχ-tensor parameters. The ―diagonal rule‖ used in (Pintacuda et al., 2004) to manually measure PCSs has also been exploited in (Schmitz et al., 2006) to automatically assign paramagnetic chemical shifts of a full 15 N-HSQC spectrum, given a known 3D structure and the list of assigned diamagnetic
1.3 Computational study of paramagnetic proteins. 13 resonances. The software Echidna was developed to overcome the difficulties of sequential assignment of cross peaks in a paramagnetic spectrum. It works as follow: Firstly, a small number n1 of paramagnetic peaks are paired with diamagnetic peaks by automatically screening unambiguous possibilities along the diagonal of a 2D 15 N-HSQC spectrum. Secondly, a Δχ-tensor is calculated to minimize the root mean square deviations between the n1 experimental and calculated PCSs. Thirdly, the Δχ-tensor is used to predict for each diamagnetic cross peak the area of the spectrum where the paramagnetic partner is expected. This area is centered on the back- calculated PCS value, and defines a much smaller zone than the diagonal strip used in the first step. More paramagnetic peaks are unambiguously assigned. The two last steps are iterated until convergence. A final assignment is performed in order to yield the overall best assignment of all cross peaks after convergence of the method. This assignment uses the Hungarian method which finds in a polynomial time the optimal assignment among the n! possibilities, with n being the number of peaks to assign. A complete flow chart of the method is given in Figure 1.9.
Page 1 and 2: Computational study of proteins wit
Page 3 and 4: iii Schmitz C, Stanton-Cook MJ, Su
Page 5 and 6: v Acknowledgements I would like to
Page 7 and 8: vii Keywords paramagnetic nmr, pseu
Page 9 and 10: ix 2.3.3 Manual resonance assignmen
Page 11 and 12: xi List of Figures Figure 1.1 NMR e
Page 13 and 14: xiii
Page 15: xv List of Abbreviations α Subunit
Page 18 and 19: 2 Chapter 1. Introduction. The scal
Page 20 and 21: 4 Chapter 1. Introduction. In contr
Page 22 and 23: 6 Chapter 1. Introduction. as the c
Page 24 and 25: 8 Chapter 1. Introduction. Figure 1
Page 26 and 27: 10 Chapter 1. Introduction. Protein
Page 30 and 31: 14 Chapter 1. Introduction. Figure
Page 32 and 33: 16 Chapter 1. Introduction. Figure
Page 34 and 35: 18 Chapter 1. Introduction. (Bugaye
Page 36 and 37: 20 Chapter 1. Introduction. The det
Page 38 and 39: 22 Chapter 1. Introduction. (ii) It
Page 40 and 41: 24 Chapter 1. Introduction. Chapter
Page 42 and 43: 26 Chapter 1. Introduction. Su XC,
Page 44 and 45: 28 Chapter 2. Possum: paramagnetica
Page 78 and 79:
62 Chapter 2. Possum: paramagnetica
Page 80 and 81:
Page 82 and 83:
Page 84 and 85:
Page 86 and 87:
Page 88 and 89:
Page 91 and 92:
Chapter 3 Numbat: new user-friendly
Page 93 and 94:
3.4 Introduction 3.4 Introduction.
Page 95 and 96:
3.5 Algorithm. 79 rhombic component
Page 97 and 98:
3.6 Program Features. 81 each fitti
Page 99 and 100:
3.6 Program Features. 83 A new PCS
Page 101 and 102:
3.6 Program Features. 85 Carlo prot
Page 103 and 104:
3.7 Study case. 87 The coordinates
Page 105 and 106:
3.7 Study case. 89 illustrates how
Page 107 and 108:
3.7 Study case. 91 can be used to a
Page 109 and 110:
3.9 Acknowledgment. 93 variables, t
Page 111 and 112:
3.10 References. 95 Galassi M, Davi
Page 113 and 114:
3.11 Supporting information. 97 Zwe
Page 115:
3.11 Supporting information. 99 Tab
Page 118 and 119:
102 Chapter 4. Protein Structure De
Page 120 and 121:
Page 122 and 123:
Page 124 and 125:
Page 126 and 127:
Page 128 and 129:
Page 130 and 131:
Page 132 and 133:
Page 134 and 135:
Page 136 and 137:
Page 138 and 139:
Page 140 and 141:
Page 142 and 143:
Page 145 and 146:
Chapter 5 Conclusion and perspectiv
Page 147 and 148:
5.1 The use of PCS for structure de
Page 149 and 150:
5.1 The use of PCS for structure de
Page 151 and 152:
5.3 The use of PCS for protein dock
show all

Thesis Title: Subtitle - NMR Spectroscopy Research Group

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?