Thesis final - after defense-7 - Jacobs University

3.1.3. Results and Discussion ............................................................................................... 45
3.1.3.1. Chromatographic profiles of different ligands .................................................... 45
3.1.3.2. Two dimensional gel electrophoresis and proteins identification ....................... 47
3.1.3.3. Influence of the ligands chemistries and average protein properties .................. 55
3.1.3.3.1. Average hydrophobicity (AH) ............................................................................. 55
3.1.3.3.2. Average polarity (AP) .......................................................................................... 59
3.1.3.3.3. Average flexibility (AF)....................................................................................... 62
3.1.3.3.4. Average Bulkiness (AB) ...................................................................................... 65
3.1.3.3.5. Average surface hydrophobicity (ASH)............................................................... 66
3.1.3.4. Comparative studies of the hydrophobic ligands based on average protein
properties .......................................................................................................................... 68
3.1.3.5. Application note .................................................................................................. 70
3.1.4. Partial conclusions ...................................................................................................... 71
3.2. Influence of the different base support chemistries during hydrophobic interaction
chromatography ........................................................................................................................ 72
3.2.1. Summary .................................................................................................................... 72
3.2.2. Chemistry of the base supports .................................................................................. 73
3.2.3. Results and Discussion ............................................................................................... 75
3.2.3.1. Chromatographic fractionation ........................................................................... 75
3.2.3.2. The electrophoretic separation and proteins identification ................................. 78
3.2.3.3. Influence of the base support chemistries and average protein properties .......... 85
3.2.3.3.1. Average hydrophobicity (AH) ............................................................................. 85
3.2.3.3.2. Average polarity (AP) .......................................................................................... 89
3.2.3.3.3. Average surface hydrophobicity (ASH)............................................................... 93
3.2.3.3.4. Other protein properties affecting the chromatographic behavior ....................... 95
3.2.3.4. Comparative studies of the different base supports based on average protein
properties .......................................................................................................................... 98
3.2.3.5. Application note ................................................................................................ 100
3.2.3. Partial conclusions .................................................................................................... 101
3.3. Protein distributions on 2-D gels: the influence of molecular weight and isoelectric point
coordinates ............................................................................................................................. 102
3.3.1. Summary .................................................................................................................. 102
3.3.2. Results and Discussion ............................................................................................. 103
3.3.2.1. Unfolding of proteins during HIC ..................................................................... 103
3