Preface	8
	Contents	11
	Symbols	15
	1 The three-dimensional structure of proteins	17
	1.1 Structure of the native state	17
	1.2 Protein folding transition states	25
	1.3 Structural determinants of the folding rate constants	28
	1.4 Support of structure determination by protein folding simulations	36
	2 Liquid chromatography of biomolecules	39
	2.1 Ion exchange chromatography	39
	2.2 Gel filtration chromatography	44
	2.3 Affinity chromatography	47
	2.4 Counter-current chromatography and ultrafiltration	49
	3 Mass spectrometry	53
	3.1 Principles of operation and types of spectrometers	53
	3.1.1 Sector mass spectrometer	54
	3.1.2 Quadrupole mass spectrometer	55
	3.1.3 Ion trap mass spectrometer	55
	3.1.4 Time-of-flight mass spectrometer	56
	3.1.5 Fourier transform mass spectrometer	59
	3.1.6 Ionization, ion transport and ion detection	60
	3.1.7 Ion fragmentation	61
	3.1.8 Combination with chromatographic methods	62
	3.2 Biophysical applications	65
	4 X-ray structural analysis	75
	4.1 Fourier transform and X-ray crystallography	75
	4.1.1 Fourier transform	75
	4.1.2 Protein X-ray crystallography	85
	4.1.2.1 Overview	85
	4.1.2.2 Production of suitable crystals	85
	4.1.2.3 Acquisition of the diffraction pattern	87
	4.1.2.4 Determination of the phases: heavy atom replacement	92
	4.1.2.5 Calculation of the electron density and refinement	99
	4.1.2.6 Cryocrystallography and time-resolved crystallography	100
	4.2 X-ray scattering	101
	4.2.1 Small angle X-ray scattering (SAXS)	101
	4.2.2 X-ray backscattering	104
	5 Protein infrared spectroscopy	107
	5.1 Spectrometers and devices	108
	5.1.1 Scanning infrared spectrometers	108
	5.1.2 Fourier transform infrared (FTIR) spectrometers	108
	5.1.3 LIDAR, optical coherence tomography, attenuated total reflection and IR microscopes	112
	5.2 Applications	118
	6 Electron microscopy	123
	6.1 Transmission electron microscope (TEM)	123
	6.1.1 General design	123
	6.1.2 Resolution	125
	6.1.3 Electron sources	126
	6.1.4 TEM grids	128
	6.1.5 Electron lenses	128
	6.1.6 Electron-sample interactions and electron spectroscopy	131
	6.1.7 Examples of biophysical applications	133
	6.2 Scanning transmission electron microscope (STEM)	134
	7 Scanning probe microscopy	137
	7.1 Atomic force microscope (AFM)	137
	7.2 Scanning tunneling microscope (STM)	149
	7.3 Scanning nearfield optical microscope (SNOM)	151
	7.3.1 Overcoming the classical limits of optics	151
	7.3.2 Design of the subwavelength aperture	154
	7.3.3 Examples of SNOM applications	158
	7.4 Scanning ion conductance microscope, scanning thermal microscope and further scanning probe microscopes	159
	8 Biophysical nanotechnology	163
	8.1 Force measurements in single protein molecules	163
	8.2 Force measurements in a single polymerase-DNA complex	166
	8.3 Molecular recognition	168
	8.4 Protein nanoarrays and protein engineering	171
	8.5 Study and manipulation of protein crystal growth	174
	8.6 Nanopipettes, molecular diodes, self-assembled nanotransistors, nanoparticle-mediated transfection and further biophysical nanotechnologies	175
	9 Proteomics: high throughput protein functional analysis	181
	9.1 Target discovery	182
	9.2 Interaction proteomics	184
	9.3 Chemical proteomics	188
	9.4 Lab-on-a-chip technology and mass-spectrometric array scanners	189
	9.5 Structural proteomics	190
	10 Ion mobility spectrometry	191
	10.1 General design of spectrometers	191
	10.2 Resolution and sensitivity	196
	10.3 IMS-based “sniffers”	199
	10.4 Design details	200
	10.5 Detection of biological agents	209
	11 ?-Value analysis	213
	11.1 The method	213
	11.2 High resolution of six protein folding transition states	215
	12 Evolutionary computer programming	219
	12.1 Reasons for the necessity of self-evolving computer programs	219
	12.2 General features of the method	219
	12.3 Protein folding and structure simulations	222
	12.4 Evolution of nanooptical devices made from nanoparticles	223
	12.4.1 Materials and methods	223
	12.4.2 Results and discussion	224
	12.5 Further potential applications	226
	13 Conclusions	229
	References	231
	Index	263
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