Publications .................................................... 5
1 Introduction ................................................ 15
1.1 Open problem 1 ......................................... 17
1.2 Open problem 2 ......................................... 19
2 Background .................................................. 23
2.1 Membrane proteins ...................................... 23
2.1.1 Biological membranes ............................ 23
2.1.2 Membrane protein classification ................. 24
2.1.3 Membrane protein structural features and amino
acid preferences ................................ 27
2.1.4 Membrane protein folding ........................ 29
2.1.5 Membrane proteins and diseases .................. 31
2.1.6 Experimental techniques for membrane protein
structure determination ......................... 33
2.1.7 Molecular Dynamics studies of membrane
proteins ........................................ 36
2.1.8 Topology prediction methods for transmembrane
proteins ........................................ 37
2.1.9 2.5D predictions of transmembrane proteins ...... 39
2.1.10 3D predictions of transmembrane proteins ........ 43
2.2 Function prediction of proteins ........................ 45
2.2.1 Functional annotation from sequence alone ....... 45
2.2.2 Elucidation of function from structure .......... 47
2.2.3 Membrane protein-specific motifs ................ 53
2.3 Outlook ................................................ 56
3 Transmembrane protein motifs ................................ 59
3.1 Introduction ........................................... 60
3.2 Results ................................................ 61
3.2.1 The algorithm for motif generation .............. 61
3.2.2 Results from the structural fragment
clustering ...................................... 63
3.2.3 Motifs and functional annotation ................ 68
3.2.4 Family-specific and cross-family motifs ......... 73
3.2.5 Sequence motifs improve transmembrane topology
prediction tools ................................ 74
3.3 Discussion and outlook ................................. 78
3.4 Materials and Methods .................................. 80
3.4.1 Dataset ......................................... 80
3.4.2 Fragments generation and description ............ 80
3.4.3 Generation of the motif library ................. 82
3.4.4 Prosite comparison by aligning regular
expressions ..................................... 83
3.4.5 Functional Annotation of motifs ................. 84
3.4.6 Statistical and biological significance of
motifs .......................................... 85
4 A scientific wiki for transmembrane protein motifs .......... 93
4.1 Motivation ............................................. 94
4.2 MotifWiki .............................................. 95
4.2.1 The MotifWiki database .......................... 95
4.2.2 The MotifWiki web-server ........................ 96
4.3 Discussion and outlook ................................ 103
5 Transmembrane protein stability investigated by Atomic
Force Microscopy ........................................... 107
5.1 Introduction .......................................... 109
5.2 Algorithm for pattern recognition of membrane-
protein unfolding pathways ............................ 111
5.2.1 Data Preparation: filtering bad curves and
determining zero-force baseline and contact
point .......................................... 112
5.2.2 Noise Reduction: singular value
decomposition .................................. 113
5.2.3 Curve alignment with dynamic programming ....... 113
5.2.4 Defining unfolding classes with hierarchical
clustering ..................................... 114
5.2.5 Peak detection ................................. 114
5.3 Progressive alignment of force spectra ................ 115
5.4 Evaluation of the pattern recognition algorithm for
SMFS unfolding data ................................... 116
5.4.1 Experimental setup ............................. 116
5.4.2 Spurious curves and peak detection ............. 117
5.4.3 Unfolding pathways ............................. 117
5.4.4 Curve alignment vs. manual annotation: 76%
success rate ................................... 117
5.4.5 Integration of the pattern recognition
algorithm into the DURIN software .............. 118
5.5 Case of study I: bacteriorhodopsin and four mutants
unfolding data ........................................ 120
5.5.1 Motivation ..................................... 120
5.5.2 Residue-residue contact area analysis .......... 121
5.5.3 Coarse-grained model of interaction energies
for amino acids ................................ 125
5.6 Case of study II: Two functional states of bovine
rhodopsin ............................................. 129
5.6.1 Motivation ..................................... 129
5.6.2 Analysis of SMFS experiments of bovine
rhodopsin ...................................... 129
5.7 Highly conserved sequence-structure motifs and
unfolding barriers .................................... 131
5.7.1 Motifs associated to unfolding barriers in
bacteriorhodopsin (PDB ID: 1brr) ............... 131
5.7.2 Motifs associated to unfolding barriers in
sodium/proton antiporter (PDB ID: 1zcd) ........ 132
5.8 Discussion and outlook ................................ 134
6 Summary and future work .................................... 139
6.1 Open problem 1 revisited .............................. 139
6.2 Open problem 2 revisited .............................. 140
6.3 Future work ........................................... 141
6.3.1 Improve classification schemes of
transmembrane proteins ......................... 141
6.3.2 Detection of distantly related transmembrane
proteins ....................................... 142
6.3.3 Creation of a tool for constrained
transmembrane topology predictions ............. 142
6.3.4 Prediction of transmembrane protein
structures constrained by experimental data .... 142
References .................................................... 147
Acknowledgements .............................................. 161
| 
