Programmed Self-assembly of Coiled-coil Peptides Público
Xu, Chunfu (2013)
Abstract
Programmed Self-assembly of Coiled-coil Peptides
By Chunfu Xu
Self-assembly is an extension of the central dogma of molecular biology, bridging the realm of linear information and the realm of protein assemblies. However, due to our limited understanding of principles of chemical self-assembly, the level of complexity of synthetic self-assembling systems pales in comparison to what nature exhibits all around us. With their relatively straightforward intermolecular interfaces, coiled coils could be a model system for studying rules of protein self-assembly and rationally designing complex peptide supramolecular assemblies. Surrounding the ubiquitous structural motif, coiled coil, different research topics will be presented including controlling peptide self-assembly through metal-induced registry shift, recoding heptameric coiled coil for nanotube formation, and designing large-diameter helical nanotubes. During these studies, various biophysical measurements conducted in solution and the solid state over multiple length scales of structural hierarchy were employed to verify our hypotheses. These studies provided design strategies for dynamically reconfigurable nanoscale materials and nano-porous protein-based materials, and investigated the primary sequence determinants which underlie the self-assembly processes of coiled coils. The results of these studies described in this volume are of both theoretical and practical significance in understanding primary rules of peptide/protein self-assembly and rational design of functional protein-based materials.
Table of Contents
Table of Contents
Chapter I: Introduction. 16
1.1 The importance of chemical self-assembly. 1
1.1.1 Self-assembly created the complex hierarchy of life. 3
1.1.2 Self-assembly is also the only practical approaches for making a variety of nanostructures. 5
1.2 Introduction to coiled coils. 8
1.3 Extended KIH packing and higher-order coiled coils. 12
1.4 Coiled-coil supramolecular assemblies. 17
1.5 Summary. 23
1.6 References: 25
Chapter II: Controlling Self-assembly of a Peptide-Based Material via Metal-Ion Induced Registry Shift 30
2.1 Introduction and Design. 30
2.2 Results and Discussion. 34
2.2.1 Circular Dichroism.. 34
2.2.2 Flow Linear Dichroism.. 36
2.2.3 Transmission Electron Microscopy. 37
2.2.4 Analytical Ultracentrifugation. 38
2.2.5 113Cd NMR spectroscopy and 111mCd PAC spectroscopy. 40
2.2.6 Non-denaturing mass spectrometry. 43
2.3 Summary. 45
2.4 Materials and methods. 46
2.5 References: 54
Chapter III: Rational Design of Helical Nanotubes from Self-assembly of Coiled-coil Lock Washers 57
3.1 Introduction. 57
3.2 Design of 7HSAP1 Sequence. 63
3.3 Results and Discussion. 65
3.3.1 Circular Dichroism and Flow Linear Dichroism.. 65
3.3.2 Electron Microscopy. 67
3.3.3 Solution X-ray scattering measurements. 73
3.3.4 X-ray Fiber Diffraction. 76
3.3.5 Solid-state NMR measurements. 78
3.3.6 Computational Modeling. 84
3.3.7 Fluorescence Spectroscopy. 90
3.3 Summary. 95
3.4 Materials and methods. 99
3.5 References. 114
Chapter IV: De Novo Design of Helical Nanotubes from Coiled-coil Peptides. 127
4.1 Introduction. 127
4.2 Barrel3CLys, a bilayered helical nanotube. 130
4.2.1 Sequence Design. 130
4.2.2 Result and Discussion. 131
4.2.2.1 Circular Dichroism and Flow Linear Dichroism.. 131
4.2.2.2 Negative-stain Electron Microscopy and Mass-per-Length Measurements. 133
4.2.2.3 Small Angle X-ray Scattering. 136
4.2.2.4 Cryo-EM Helical Reconstruction. 139
4.2.2.5 D-Barrel3CLys, a mirror image of Barrel3CLys. 142
4.3 Barrel3CArg, a conservative mutant of Barrel3CLys. 145
4.3.1 Sequence of Barrel3CArg. 145
4.3.2 Results and Discussion. 146
4.3.2.1 Circular Dichroism and Flow Linear Dichroism.. 146
4.3.2.2 Negative-stain Electron Microscopy. 148
4.3.2.3 Solution X-ray Measurements. 149
4.3.2.4 Cryo-EM Helical Reconstruction. 152
4.3.2.5 Computational Modeling. 156
4.3.2.6 Arginine 13, the structural switch. 159
4.4 Summary. 162
4.5 Materials and methods. 163
Chapter V: Conclusion and outlook. 176
5.1 Conclusion. 176
5.2 Outlook. 178
5.3 References. 180
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