Methods in Modern Biophysics, 2/e

Description

Incorporating dramatic recent advances, "Methods in Modern Biophysics" presents a fresh and timely introduction to modern biophysical methods. This innovative text surveys and explains the ten key biophysical methods, including those related to biophysical nanotechnology, scanning probe microscopy, X-ray crystallography, ion mobility spectrometry, mass spectrometry, and proteomics. Containing much information previously unavailable in tutorial form, "Methods in Modern Biophysics" employs worked examples and more than 260 illustrations to fully detail the techniques and their underlying mechanisms. The book was written for advanced undergraduate and graduate students, postdocs, researchers, lecturers and professors in biophysics, biochemistry, general biology and related fields.

Table of contents

1 The three-dimensional structure of proteins ................................................. 1
1.1 Structure of the native state ................................................................. 1
1.2 Protein folding transition states ............................................................ 9
1.3 Structural determinants of the folding rate constants ......................... 12
1.4 Support of structure determination by protein folding simulations ......... 20
2 Liquid chromatography of biomolecules ................................................... 23
2.1 Ion exchange chromatography .............................................................. 23
2.2 Gel filtration chromatography .............................................................. 28
2.3 Affinity chromatography ...................................................................... 31
2.4 Counter-current chromatography and ultrafiltration ................................ 33
3 Mass spectrometry ................................................................................. 37
3.1 Principles of operation and types of spectrometers ................................. 37
3.1.1 Sector mass spectrometer ................................................................. 38
3.1.2 Quadrupole mass spectrometer ........................................................ 39
3.1.3 Ion trap mass spectrometer .............................................................. 39
3.1.4 Time-of-flight mass spectrometer .................................................... 40
3.1.5 Fourier transform mass spectrometer ............................................... 43
3.1.6 Ionization, ion transport and ion detection ...................................... 44
3.1.7 Ion fragmentation ............................................................................. 45
3.1.8 Combination with chromatographic methods .................................. 46
3.2 Biophysical applications ...................................................................... 49
4 X-ray structural analysis ....................................................................... 59
4.1 Fourier transform and X-ray crystallography........................................... 59
4.1.1 Fourier transform ............................................................................. 59
4.1.2 Protein X-ray crystallography .......................................................... 69
4.1.2.1 Overview .................................................................................. 69
4.1.2.2 Production of suitable crystals .................................................. 69
4.1.2.3 Acquisition of the diffraction pattern ........................................ 71
XII Contents
4.1.2.4 Determination of the phases: heavy atom replacement ............. 76
4.1.2.5 Calculation of the electron density and refinement .................. 83
4.1.2.6 Cryocrystallography and time-resolved crystallography........... 84
4.2 X-ray scattering .......................................................................... 85
4.2.1 Small angle X-ray scattering (SAXS) ............................................. 85
4.2.2 X-ray backscattering ............................................................. 88
5 Protein infrared spectroscopy ............................................................ 91
5.1 Spectrometers and devices ............................................................ 92
5.1.1 Scanning infrared spectrometers ................................................... 92
5.1.2 Fourier transform infrared (FTIR) spectrometers ............................ 92
5.1.3 LIDAR, optical coherence tomography, attenuated total
reflection and IR microscopes
................. 96
5.2 Applications .......................................................................... 102
6 Electron microscopy .................................................................. 107
6.1 Transmission electron microscope (TEM)...................................... 107
6.1.1 General design ..................................................................... 107
6.1.2 Resolution ........................................................................... 109
6.1.3 Electron sources ................................................................... 110
6.1.4 TEM grids ............................................................................. 112
6.1.5 Electron lenses ....................................................................... 112
6.1.6 Electron-sample interactions and electron spectroscopy ................ 115
6.1.7 Examples of biophysical applications ........................................... 117
6.2 Scanning transmission electron microscope (STEM) ............................ 118
7 Scanning probe microscopy .............................................................. 121
7.1 Atomic force microscope (AFM) .......................................................... 121
7.2 Scanning tunneling microscope (STM) ................................................. 133
7.3 Scanning nearfield optical microscope (SNOM) ................................... 135
7.3.1 Overcoming the classical limits of optics ...................................... 135
7.3.2 Design of the subwavelength aperture ........................................... 138
7.3.3 Examples of SNOM applications ................................................... 142
7.4 Scanning ion conductance microscope, scanning thermal
microscope and further scanning probe microscopes
.................... 143
8 Biophysical nanotechnology ............................................................... 147
8.1 Force measurements in single protein molecules .................................. 147
8.2 Force measurements in a single polymerase-DNA complex .................. 150
Contents XIII
8.3 Molecular recognition ........................................................................... 152
8.4 Protein nanoarrays and protein engineering .......................................... 155
8.5 Study and manipulation of protein crystal growth ................................. 158
8.6 Nanopipettes, molecular diodes, self-assembled nanotransistors,
nanoparticle-mediated transfection and further biophysical
nanotechnologies
......... 159
9 Proteomics: high throughput protein functional analysis ....................... 165
9.1 Target discovery .......................................................................... 166
9.2 Interaction proteomics ................................................................. 168
9.3 Chemical proteomics ................................................................... 172
9.4 Lab-on-a-chip technology and mass-spectrometric array scanners ....... 173
9.5 Structural proteomics ................................................................... 174
10 Ion mobility spectrometry .................................................................... 175
10.1 General design of spectrometers ......................................................... 175
10.2 Resolution and sensitivity ................................................................ 180
10.3 IMS-based “sniffers” ...................................................................... 183
10.4 Design details ................................................................................. 184
10.5 Detection of biological agents ......................................................... 193
11 Φ-Value analysis ............................................................................... 197
11.1 The method ..................................................................................... 197
11.2 High resolution of six protein folding transition states ....................... 199
12 Evolutionary computer programming ......................................................... 203
12.1 Reasons for the necessity of self-evolving computer programs .......... 203
12.2 General features of the method ........................................................... 203
12.3 Protein folding and structure simulations ........................................... 206
12.4 Evolution of nanooptical devices made from nanoparticles .............. 207
12.4.1 Materials and methods ................................................................ 207
12.4.2 Results and discussion ................................................................ 208
12.5 Further potential applications .............................................................. 210
13 Conclusions .................................................................................... 213
References ........................................................................................ 215
Index ................................................................................................ 247

《現代生物物理學方法》是一本介紹現代生物物理學方法的新鮮及及時的教材，包含了最新的進展。這本創新的教材介紹了十種關鍵的生物物理學方法，包括與生物物理納米技術、掃描探針顯微術、X射線晶體學、離子遷移光譜法、質譜法和蛋白質組學相關的方法。《現代生物物理學方法》提供了許多以前無法在教學形式中獲得的資訊，並通過實例和260多幅插圖詳細介紹了這些技術及其基本原理。這本書適用於高年級本科生、研究生、博士後、研究人員、講師和教授，以及生物物理學、生物化學、一般生物學和相關領域的專業人士。

目錄:
1. 蛋白質的三維結構................................................. 1
1.1 原生態結構................................................................. 1
1.2 蛋白質折疊過渡態............................................................ 9
1.3 折疊速率常數的結構決定因素............................................ 12
1.4 蛋白質折疊模擬對結構確定的支持.................................... 20
2. 生物分子的液相色譜................................................... 23
2.1 離子交換色譜.............................................................. 23
2.2 凝膠過濾色譜.............................................................. 28
2.3 亲和色譜...................................................................... 31
2.4 逆流色譜和超濾........................................................ 33
3. 質譜法................................................................................. 37
3.1 操作原理和質譜儀類型................................................. 37
3.1.1 扇形質譜儀................................................................. 38
3.1.2 四極質譜儀........................................................... 39
3.1.3 離子陷阱質譜儀.............................................................. 39
3.1.4 飛行時間質譜儀.................................................... 40
3.1.5 傅立葉變換質譜儀............................................... 43
3.1.6 離子化、離子傳輸和離子檢測...................................... 44
3.1.7 離子碎裂..................................................................... 45
3.1.8 與色譜法結合.......................................................... 46
3.2 生物物理學應用.............................................................. 49
4. X射線結構分析............................................................... 59
4.1 傅立葉變換和X射線晶體學........................................... 59
4.1.1 傅立葉變換................................................................. 59
4.1.2 蛋白質X射線晶體學.................................................