Calcium-Binding Protein Protocols Calcium-Binding Protein Protocols

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Isotope Labeling 265 and 15 N labeled ingredients, then carefully pour this back into the flask containing 13 C and 15 N medium in 99.9% D 2O. 25. Because of the higher level of Mg 2+ and Fe 2+ salts used here and their lower solubility in D 2O than in H 2O, visibly high levels of precipitates may form during incubations. These are mainly metal hydroxides. These precipitates have no detrimental effect on cell growth, but they scatter visible light as cells do. Thus, the precipitates may cause a problem in the measurement of cell growth (typically monitored by measuring OD 600nm). However, as the cells approach mid-log phase, the precipitates clear away when the minerals are consumed by bacteria, OD 600nm becomes a reliable indicator for cell growth. 26. Because cell metabolism is slower in D 2O, induction will be longer, perhaps 3–4 times longer than in H 2O. Accurate induction times should be determined by time-courses. 27. This method is good to recycle D 2O used previously to grow cells and/or D 2O accumulated over the years from various NMR samples. A good rule of thumb is: distill until visibly clear, then distill one more time. This distillate will still contain trace amounts of volatile organic compounds, which are toxic to biological organizms so will not support life. It can therefore be stored indefinitely in the refrigerator or at room temperature. 28. This step should be done immediately before use. Because this D 2O will now support life, it should be stored at refrigeration temperature otherwise mould, alga, and other exotica will grow in it and it will have to be recycled again. 29. The most convenient way to determine the percent deuteration of the D 2O recycled is gravimetrically. References 1. Wüthrich, K. (1986) NMR of proteins and nucleic acids. Wiley, New York. 2. Cavanagh, J., Fairbrother, W. J., Palmer, A. G., III, and Skelton, N. J. (1996) <strong>Protein</strong> NMR spectroscopy: Principles and Practice. Academic, San Diego, California. 3. Gardner, K. H. and Kay, L. E. (1998) The use of 2H, 13C, 15N multidimensional NMR to study the structure and dynamics of proteins. Annu. Rev. Biophys. Biomol. Struct. 27, 357–406. 4. Venters, R. A., Calderone, T. L., Spicer, L. D., and Fierke, C. A. (1991) Uniform 13C isotope labeling of proteins with sodium acetate for NMR studies: application to human carbonic anhydrase II. Biochemistry 30, 4491–4494. 5. Studier, F. W., Rosenberg, A. H., Dunn, J. J., and Dubendorff, J. W. (1990) Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 185, 60–89. 6. Gagne, S. M., Li, M. X., McKay, R. T., and Sykes, B. D. (1998) The NMR angle on troponin C. Biochem. Cell Biol. 76, 302–312. 7. Muchmore, D. C., McIntosh, L. P., Russell, C. B., Anderson, D. E., and Dahlquist, F. W. (1989) Expression and nitrogen-15 labeling of proteins for proton and nitrogen-15 nuclear magnetic resonance. Methods Enzymol. 177, 44–73.
20 Dean, Kelsey, and Reik
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Methods in Molecular BiologyTM Biol
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M E T H O D S I N M O L E C U L A R
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© 2002 Humana Press Inc. 999 River
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Preface Calcium plays an important
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Contents Dedication ...............
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Contents xi 26 Enzymatic Assays to
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xiv Contents of Companion Volume 15
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xvi Contributors LESLIE D. HICKS
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20 Dean, Kelsey, and Re
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2 Yazawa
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4 Yazawa Fig. 1. Schematic represen
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6 Yazawa Fig. 2. Shop drawing for t
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8 Yazawa DuPont-NEN and the molar c
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10 Yazawa Fig. 4. Examples of the C
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12 Yazawa 5. Plot the calculated Ca
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14 Yazawa 12. Starovasnik, M. A., D
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16 Fig. 1. Molecular structures and
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18 Linse 7. 0.1 M EDTA. Dissolve 37
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20 Linse iterate the other paramete
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22 Linse tive to small alterations
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24 Linse References 1. Tsien, R. Y.
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26 Haiech and Kilhoffer to use the
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28 Haiech and Kilhoffer where γ is
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30 Haiech and Kilhoffer reporter gr
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32 Haiech and Kilhoffer tein with i
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34 Haiech and Kilhoffer Calmodulin
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36 Haiech and Kilhoffer Fig. 3. Mec
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38 Haiech and Kilhoffer We may sugg
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40 Haiech and Kilhoffer 29. Stewart
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42 Haiech and Kilhoffer 62. Becking
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44 Martin and Bayley Fig. 1. Absorp
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46 Martin and Bayley evaporation. C
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48 Martin and Bayley 4. Set the tem
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50 Martin and Bayley Fig. 2. Near-U
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52 Martin and Bayley discussed by M
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54 Martin and Bayley References 1.
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58 Fabian and Vogel are equipped wi
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60 Fabian and Vogel The penetration
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62 Fabian and Vogel perature, ionic
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64 Fabian and Vogel Fig. 3. IR spec
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66 Fabian and Vogel Fig. 4. Lower t
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68 Fabian and Vogel These correlati
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70 Fabian and Vogel Fig. 5. Amide I
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72 Fabian and Vogel 4. ATR-FTIR spe
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74 Fabian and Vogel 25. Georg, H.,
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76 Weljie and Vogel Fig. 1. Simplif
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78 Weljie and Vogel Fig. 3. Steady-
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80 Weljie and Vogel VIS spectrophot
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82 Weljie and Vogel 4. A series of
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84 Table 1 Advanced Fluorescence Me
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86 Weljie and Vogel References 1. L
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90 Johnson and Tikunova is removed
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92 Johnson and Tikunova function of
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94 Johnson and Tikunova Fig. 2. Rat
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96 Johnson and Tikunova Fig. 3. Rat
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98 Johnson and Tikunova Fig. 4. Ca
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100 Johnson and Tikunova cence inte
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102 Johnson and Tikunova 6. Johnson
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104 Julenius Fig. 1. Under conditio
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106 Julenius 3. Mix 50 µL of NHS s
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108 Julenius Fig. 3. A typical sens
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110 Julenius (2), is used in the Ia
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114 Lopez and Makhatadze Fig. 1. Th
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116 Lopez and Makhatadze 6. Buffers
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118 Lopez and Makhatadze The excess
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122 Lopez and Makhatadze Fig. 1. Is
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124 Lopez and Makhatadze 2.5 mL are
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126 Lopez and Makhatadze pendence o
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128 Hicks et al. equipped with a pu
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130 Hicks et al. Fig. 2. Debye plot
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132 Hicks et al. and 1.0 mg/mL for
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134 Hicks et al. Fig. 3. Sedimentat
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136 Hicks et al. 5. Hayes, D. B. (M
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138 Trewhella and Krueger This chap
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140 Trewhella and Krueger of the sc
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142 Trewhella and Krueger Table 1 C
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144 Trewhella and Krueger beam, or
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146 Trewhella and Krueger At very l
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148 Trewhella and Krueger analytica
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150 Trewhella and Krueger collapsed
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152 Trewhella and Krueger P(r) func
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154 Trewhella and Krueger Fig. 3. S
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156 Trewhella and Krueger concentra
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158 Trewhella and Krueger by a Nati
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162 Doherty-Kirby and Lajoie metal
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164 Doherty-Kirby and Lajoie two Ca
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166 Doherty-Kirby and Lajoie 7. Pep
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168 Doherty-Kirby and Lajoie Fig. 1
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170 Doherty-Kirby and Lajoie Fig. 2
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172 Doherty-Kirby and Lajoie 5. Alt
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174 Doherty-Kirby and Lajoie phosph
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176 Shaw Fig. 1. Ribbon drawing of
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178 Shaw 2.3. Other 1. Chemicals fo
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180 Shaw 3.3.3. Purification 1. Con
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182 Shaw 10. Hodges, R. S., Semchuk
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184 Brokx and Vogel Table 1 An Over
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186 Brokx and Vogel Fig. 1. UV abso
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188 Brokx and Vogel Fig. 2. 20% SDS
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190 Brokx and Vogel it through an a
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192 Brokx and Vogel 8. Weljie, A. M
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196 Berliner Fig. 1. Aqueous X-band
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198 Berliner in the Bruker instrume
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200 Berliner Fig. 3. ESR spectra of
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202 Berliner Fig. 5. Low-temperatur
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204 Berliner 14. Musci, G., Reed, G
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206 Clarke and Vogel cal calcium-bi
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208 Clarke and Vogel Fig. 1. 113 Cd
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Page 233 and 234: 214 Clarke and Vogel The linewidth
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Page 237 and 238: 218 Drakenberg sands, of Hertz broa
Page 239 and 240: 220 Drakenberg 1. Bloch equations m
Page 241 and 242: 222 Drakenberg Fig. 2. Measurement
Page 243 and 244: 224 Drakenberg Fig. 3. (A) 43 Ca NM
Page 245 and 246: 226 Drakenberg Fig. 5. 43 Ca NMR sp
Page 247 and 248: 228 Drakenberg NMR at sub-mM concen
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Page 251 and 252: 232 Weljie and Heringa Table 1 Amin
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Page 267 and 268: 248 Weljie and Heringa ment require
Page 269 and 270: 250 Weljie and Heringa 10. Kawasaki
Page 271 and 272: 252 Weljie and Heringa 44. Thompson
Page 273 and 274: 254 Weljie and Heringa
Page 275 and 276: 256 Li et al. In order for these ex
Page 277 and 278: 258 Li et al. 7. Mineral Mixture (s
Page 279 and 280: 260 Li et al. 3. When the 45% D 2O/
Page 281 and 282: 262 Li et al. sion system works eff
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Page 287 and 288: 268 Mal et al. NMR data, X-PLOR (11
Page 289 and 290: 270 Mal et al. Table 1 Simulated An
Page 291 and 292: 272 Mal et al. 3.1.2.1. AMBIGUOUS D
Page 293 and 294: 274 Mal et al. molecular dynamics a
Page 295 and 296: 276 Mal et al. Assuming a rigid bod
Page 297 and 298: 278 Mal et al. assign (segid A and
Page 299 and 300: 280 Mal et al. References 1. Drenth
Page 301 and 302: 282 Mal et al. 33. Jeener, J., Meie
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Page 305 and 306: 286 Werner et al. Our research has
Page 307 and 308: 288 Fig. 1. (A) T 1, T 2 and the he
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Page 315 and 316: 296 Werner et al. Table 2 Models of
Page 317 and 318: 298 Werner et al. 9. Reinhardt, D.
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Page 321 and 322: 302 Boyd et al. utilize residual di
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Page 329 and 330: 310 Boyd et al. total and NOE energ
Page 331 and 332: 312 Boyd et al. 2. When studying mu
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316 Boyd et al. 35. Schulte-Herbrug
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318 Yap et al. image representation
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320 Yap et al. modified EF-hand is
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322 Table 1 Angle and Distance Outp
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324 Yap et al. 2. Ikura, M. (1995)
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326 Kobayashi that S-100A1 and S-10
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328 Kobayashi 3.2. Coupling of Crom
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330 Kobayashi Fig. 2. Tricine/SDS/P
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332 Kobayashi 0.1% TFA at a flow ra
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334 Kobayashi Both recombinant S-10
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336 Kobayashi Fig. 6. Affinity chro
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340 Walsh et al. verts CaM from an
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342 Walsh et al. 11. Bovine serum a
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344 Walsh et al. Fig. 1. CaM-depend
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348 Walsh et al. 3.5. NOS Reduction
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354 Walsh et al. 3. Cho, M. J., Vag
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356 Hughes et al. The electroporati
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358 Hughes et al. 4. 1 M Na 2CO 3.
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360 Hughes et al. range, repeat ste
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362 Hughes et al. Table 1 Examples
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366 Persechini of the different CaM
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368 Persechini Fig. 1. Schematic re
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370 Persechini 2. 1 L of Terrific b
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372 Persechini Fig. 4. Titration wi
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374 Persechini Table 1 Parameters f
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376 Persechini Fig. 6. The [Ca 2+ -
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378 Persechini determined if the di
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380 Persechini relatively insensiti
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382 Persechini 18. Chafouleas, J. G
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384 Török et al. actions in the c
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386 Török et al. 3. The reaction
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388 Török et al. Fig. 2. Electros
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390 Török et al. Fig. 3. Peptides
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392 Török et al. Table 1 Peptide
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394 Török et al. Fig. 7. Nanospra
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396 Török et al. Fig. 9. Nanospra
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398 Török et al. Fig. 11. Electro
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400 Török et al. 3 µM FL-calmodu
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402 Török et al. Fig. 14. Localiz
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404 Török et al. Fig. 16. Indicat
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406 Török et al. the significantl
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408 Török et al.
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410 Index substitutes, see Fluoresc
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412 Index Stern-Volmer plot, 78, 81
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414 Index Phenothiazines, see Calmo
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