Preface ........................................................ xi
Part 1 Basics
Chapter 1 Introduction ......................................... 3
1.1 Overview of biomolecular EPR spectroscopy .................. 3
1.2 How to use this book and associated software ............... 4
1.3 A brief history of bioEPR .................................. 5
Chapter 2 The Spectrometer ..................................... 9
2.1 The concept of magnetic resonance .......................... 9
2.2 The microwave frequency ................................... 12
2.3 Overview of the spectrometer .............................. 15
2.4 The resonator ............................................. 17
2.5 From source to detector ................................... 20
2.6 The magnet ................................................ 22
2.7 Phase-sensitive detection ................................. 23
2.8 Tuning the spectrometer ................................... 25
2.9 Indicative budget considerations .......................... 27
Chapter 3 The Sample .......................................... 33
3.1 Sample tube and sample size ............................... 33
3.2 Freezing and thawing ...................................... 36
3.3 Solid air problem ......................................... 39
3.4 Biological relevance of a frozen sample ................... 40
3.5 Sample preparation on the vacuum/gas manifold ............. 43
3.6 Choice of reactant ........................................ 47
3.7 Gaseous substrates ........................................ 49
3.8 Liquid samples ............................................ 50
3.9 Notes on safety ........................................... 51
Chapter 4 Experimental Key Parameters ......................... 53
4.1 Boltzmann and Heisenberg dictate optimal (P, T) pairs ..... 53
4.2 Homogeneous versus inhomogeneous lines .................... 58
4.3 Spin multiplicity and its practical implications .......... 61
Chapter 5 Resonance Condition ................................. 67
5.1 Main players in EPR theory: B, S, and I ................... 67
5.2 Anisotropy ................................................ 71
5.3 Hyperfine interactions .................................... 75
5.4 Second-order effects ...................................... 78
5.5 Low-symmetry effects ...................................... 80
5.6 Zero-field interactions ................................... 82
5.7 Integer spins ............................................. 87
5.8 Interpretation of g, A, D ................................. 89
Chapter 6 Analysis ............................................ 95
6.1 Intensity ................................................. 95
6.2 Quantification ............................................ 96
6.3 Walking the unit sphere .................................. 100
6.4 Difference spectra ....................................... 103
Part 2 Theory
Chapter 7 Energy Matrices .................................... 109
7.1 Preamble to Part 2 ....................................... 109
7.2 Molecular Hamiltonian and spin Hamiltonian ............... 112
7.3 Simple example: S = 1/2 .................................. 115
7.4 Not-so-simple example: S = 3/2 ........................... 119
7.5 Challenging example: integer spin S ≥ 2 .................. 123
7.6 Compounded (or product) spin wavefunctions ............... 131
Chapter 8 Biological Spin Hamiltonians ....................... 135
8.1 Higher powers of spin operators .......................... 135
8.2 Tensor noncolinearity .................................... 140
8.3 General EPR intensity expression ......................... 141
8.4 Numerical implementation of diagonalization solutions .... 145
8.5 A brief on perturbation theory ........................... 147
Chapter 9 Conformational Distributions ....................... 153
9.1 Classical models of anisotropic linewidth ................ 153
9.2 Statistical theory of g-strain ........................... 157
9.3 Special case of full correlation ......................... 159
9.4 A (bio)molecular interpretation of g-strain .............. 162
9.5 A-strain and D-strain: coupling to other interactions .... 164
Part 3 Specific Experiments
Chapter 10 Aqueous Solutions .................................. 169
10.1 Spin traps .............................................. 169
10.2 Spin labels in isotropic media .......................... 171
10.3 Spin labels in anisotropic media ........................ 177
10.4 Metalloproteins in solution ............................. 179
Chapter 11 Interactions ....................................... 181
11.1 Dipole-dipole interactions .............................. 181
11.2 Dipolar interaction in multicenter proteins ............. 184
11.3 Exchange interactions ................................... 188
11.4 Spin ladders ............................................ 193
11.5 Valence isomers ......................................... 196
11.6 Superparamagnetism ...................................... 197
Chapter 12 High Spins Revisited ............................... 199
12.1 Rhombograms for S = 7/2 and S = 9/2 ..................... 199
12.2 D-strain modeled as a rhombicity distribution ........... 204
12.3 Population of half-integer spin multiplets .............. 205
12.4 Intermediate-field case for S = 5/2 ..................... 207
12.5 Analytical lineshapes for integer spins ................. 208
Chapter 13 Black Box Experiments .............................. 213
13.1 EPR-monitored binding experiments ....................... 214
13.2 EPR monitoring of redox states .......................... 215
13.3 EPR monitored kinetics .................................. 221
13.4 EPR of whole cells and organelles ....................... 222
Chapter 14 Strategic Considerations ........................... 225
14.1 Bio-integrated bioEPR ................................... 225
14.2 To be advanced or not to be advanced .................... 226
14.3 Friday afternoon experiment ............................. 228
References .................................................... 231
Index ......................................................... 241
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