Thesis final - after defense-7 - Jacobs University

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Chapter 3 3.2.3.3. Influence of the base support chemistries and average protein properties 3.2.3.3.1. Average hydrophobicity (AH) Average hydrophobicity is the classical parameter based on the overall hydrophobic residues of a protein. It was reported before that in addition to hydrophobic surface residues, it is important to have a higher overall number of hydrophobic residues in the protein sequence for the stronger binding to the HIC adsorbents (62). However, the average hydrophobicity has been never investigated with the process proteomics approach. Hence, the three different hydrophobicity scales were used to calculate average hydrophobicity from the primary structures of the tabulated proteins (Tables 12-14). A statistically significant (r 2 =0.86, p < 0.0001) linear relationship was reported between the average hydrophobicity and protein retention behavior on different base supports (Figure 24 A-C). In some cases, the proteins were reported to have similar average hydrophobicity values but differed with average surface hydrophobicity. This revealed that two proteins can have the same amount of total hydrophobic residues in the primary sequence but can differ with surface hydrophobic residues. The average hydrophobicity values for some proteins were similar; however their elution position was different from each other. In this case, the proteins with high ASH retained longer than the one with low ASH, although they have similar average hydrophobicity. The proteins such as B3-TP, and F1-TP retained longer during chromatography than A4-TP and E3-TP, respectively. These results confirmed that ASH has a decisive role in protein retention during chromatography. The overall values of average hydrophobicity revealed that the scale of Miyazawa-Jernigan was differed with 1 and 2 digits to the scales of Cowan-Whittaker and Tanford, respectively. The reason for this difference could be the various direct and indirect approaches, while determining the hydrophobicity indices for individual amino acids (25, 92, 95). The mentioned scales were calculated with quite different approaches and often correlated very poorly (124). The global averages of the 85
Chapter 3 r 2 values from the data of the three base supports were 0.90, 0.89 and 0.79 for the scales of Miyazawa-Jernigan, Cowan-Whittaker and Tanford, respectively. It was also reported before that the scales of Cowan-Whittaker and Miyazawa-Jernigan have provided the best correlation for the prediction purposes in comparison to other scales (114). Although ASH is the most precise method, but the trends observed in case of average hydrophobicity cannot be ignored. The reason for this could be that, although amino acids on the surface are deciding for retention, the unfolding of proteins may occur during chromatographic experiments resulting in retention based on the total hydrophobicity of a protein. Figure 24 A: Represents the average hydrophobicity calculated by the scale of Cowan- Whittaker. The protein average hydrophobicity values of the three base supports were correlated with the respective salt concentrations where the fractions were collected. The error bars represent the standard deviations of the average hydrophobicity values in each fraction. The statistical analysis revealed a significant correlation (r 2 = 0.89; p < 0.0001). 86
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A proteome wide approach to underst
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I also want to mention the efforts
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Dedication To my father Haji Sher A
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Table of Contents 1. Introduction .
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3.3.2.2. Protein distributions on 2
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Chapter 1 1. Introduction 1.1. Intr
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Chapter 1 cell proteome to facilita
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Chapter 1 proteins. It is this diff
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Chapter 1 Salt conc. decreasing (Gr
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Chapter 1 The ions at the start of
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Chapter 1 exposure of hydrophobic r
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Chapter 1 based on the surface hydr
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Chapter 1 1.5. Analysis of the yeas
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Chapter 1 molecular weight impuriti
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Chapter 1 followed by its introduct
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Chapter 1 protein towards purificat
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Chapter 2 EXPERIMENTAL APPROACH
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Chapter 2 Yeast cell proteome and c
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Chapter 2 ligands (Toyopearl-Hexyl,
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Chapter 2 number of spots was count
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Chapter 2 (http://www.matrixscience
Page 43 and 44: Chapter 3 RESULTS AND DISCUSSION
Page 45 and 46: Chapter 3 to the retention behavior
Page 47 and 48: Chapter 3 investigate the hydrophob
Page 49 and 50: Chapter 3 the interior of the prote
Page 51 and 52: Chapter 3 A B C Figure 9: The cryst
Page 53 and 54: Chapter 3 of the comparative analys
Page 55 and 56: Chapter 3 Figure 11: Represent the
Page 57 and 58: Chapter 3 Ether, some 2-D gels were
Page 59 and 60: Chapter 3 Figure 13: Represent 2-D
Page 61 and 62: Chapter 3 Table 5. List of the iden
Page 63 and 64: Chapter 3 Table 7. List of the iden
Page 65 and 66: Chapter 3 amino acids by Cowan-Whit
Page 67 and 68: Chapter 3 Figure 15 C: Represents t
Page 69 and 70: Chapter 3 Figure 16 A: Represents t
Page 71 and 72: Chapter 3 The polarity value for th
Page 73 and 74: Chapter 3 Figure 17: Represents the
Page 77 and 78: Chapter 3 published papers with mod
Page 79 and 80: Chapter 3 ligands revealed less dif
Page 81 and 82: Chapter 3 3.2. Influence of the dif
Page 83 and 84: Chapter 3 Table 10: The chemistry o
Page 85 and 86: Chapter 3 where approximately 50% o
Page 87 and 88: Chapter 3 3.2.3.2. The electrophore
Page 89 and 90: Chapter 3 Figure 22: Represent 2-D
Page 91 and 92: Chapter 3 Table 12. List of the ide
Page 93: Chapter 3 Table 14. List of the ide
Page 97 and 98: Chapter 3 Figure 24 C: Represents t
Page 99 and 100: Chapter 3 approximately 47%. The me
Page 101 and 102: Chapter 3 Figure 25 B: Represents t
Page 103 and 104: Chapter 3 values have been reported
Page 105 and 106: Chapter 3 towards conformational ch
Page 109 and 110: Chapter 3 among the adsorbents were
Page 111 and 112: Chapter 3 3.3. Protein distribution
Page 113 and 114: Chapter 3 3.3.2.2. Protein distribu
Page 115 and 116: Chapter 3 Figure 29 B: Represent th
Page 117 and 118: Chapter 3 When the number of smalle
Page 119 and 120: Chapter 3 and hydrophobic interacti
Page 121 and 122: Chapter 4 CONCLUSIONS
Page 123 and 124: Chapter 4 • The base supports rev
Page 125 and 126: Chapter 4 additional understanding
Page 127 and 128: References 13. Lienqueo, M.E.; Lese
Page 129 and 130: References 32. Salgado, J.C.; Rapap
Page 131 and 132: References 52. B.H.J., H. Accessibl
Page 133 and 134: References 71. Xindu, G. and Regnie
Page 135 and 136: References 91. Alonso-Orgaz, S.; Mo
Page 137 and 138: References 112. Gu, Z. and Glatz, C
Page 139: References 132. Reubsaet, J.L.E. an
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Thesis final - after defense-7 - Jacobs University

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