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Chapter 4 (a) Dilutions 10 -1 10 -2 10 -3 10 -4 10 -5 10 -6 10 -7 1 2 3 4 (b) 1 2 3 4 95 kDa 72 kDa 55 kDa Fig 4.8: Complementation and expression of MQO. (a) Lack of complementation of MQO in MGM100 at 30°C in the presence of 0.2% glucose and 0.1 mM IPTG. 1- MGM100 pTrc-GroES-MQO, 2- MGM100 pTrc- GroES-GroEL, 3- MGM100 pTrc-GroES-Cpn60.2, 4- MGM100 pTrc99a. (b) SDS-PAGE gel showing minimal over expression of MQO. Samples were taken from cultures after 4 hours incubation with 0.1 mM IPTG and 0.2% arabinose. Lane 1- Prestained protein marker (Fermentas), Lane 2- MGM100 pTrc-GroES-MQO; Lane 3: MGM100 pTrc99a, Lane 4: MGM100 pTrc-GroES-GroEL. 167
Chapter 4 1 2 800 kDa 480 kDa 146 kDa 66 kDa Fig 4.9: Native gel of MQO. Native samples of MQO were taken from cells grown in LB with 0.2% glucose and induced with 0.1 mM IPTG for 4 hours. The Cpn60.2 native sample was from a purified stock. Lanes are described below: Lane 1: GroES + Cpn60.2 Lane 2: GroES + MQO 168
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FUNCTIONAL AND STRUCTURAL CHARACTER
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Abstract Proteins are involved in a
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Dedicated to Mum, Dad, and Sharique
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Index of contents Chapter 1: Introd
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2.4.5: Bacteriophage P1 Transductio
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4.1.5: Chimeras DHF & GEI 162 4.2:
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List of illustrations Chapter 1: In
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Fig 4.1: Schematic representation o
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List of tables Chapter 1: Introduct
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SDW STPKs TAE TEMED TBS Sterile dis
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Chapter 1 1.1 Protein Folding The g
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Chapter 1 Fig 1.1: The energy lands
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Chapter 1 To try and escape from th
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Chapter 1 Fig 1.2: Pathways regulat
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Chapter 1 As protein folding in Myc
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Chapter 1 1.4 The molecular chapero
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Chapter 1 1.4.1 Classes of molecula
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Chapter 1 Hsp40 DnaJ CbpA DjlA Ydj1
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Chapter 1 obtained. These revertant
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Chapter 1 1.4.3.1 The ribosome asso
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Chapter 1 (A) (B) Fig 1.7: Structur
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Chapter 1 complete the reaction cyc
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Chapter 1 1.4.3.4 The small Hsps On
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Chapter 1 1.4.3.5 Hsp90 family of c
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Chapter 1 Fig 1.11: Structure of Ht
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Chapter 1 1.4.3.6 Hsp100 family of
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Chapter 1 Fig 1.13: The different r
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Chapter 1 encoded by more than one
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Chapter 1 Fig 1.15: The illustratio
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Chapter 1 (Rommelaere et al., 1993,
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Chapter 1 Fig 1.16: Octameric struc
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Chapter 1 main difference between t
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Chapter 1 1.5.3.3 Prefoldin It has
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Chapter 1 result the substrate prot
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Chapter 1 Fig 1.18: The structures
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Chapter 1 6- If the released peptid
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Chapter 1 1.5.4.3 GroEL substrates
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Chapter 1 Recent work has suggested
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Chapter 1 (A) cpn10 cpn60.1 cpn60.2
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Chapter 1 1998). Cpn60.1 however ha
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Chapter 1 1.6 Aims of the project T
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Chapter 2 Materials and Methods: 2.
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Chapter 2 pET-cpn10- cpn60.1 Deriva
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Chapter 2 2.3 Primers: Below is a l
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Chapter 2 Mut EL.3 Rev groEL 1401-1
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Chapter 2 29R-EL-60.2 groEL 432-412
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Chapter 2 Biofilm base media: 13.6g
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Chapter 2 Preparation of bacterioph
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Chapter 2 then resuspended in 200µ
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Chapter 2 - 10X BSA (depending on t
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Chapter 2 - Colony/culture 1μl - S
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Chapter 2 The PCR cycling parameter
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Chapter 2 tubes were placed on ice
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Chapter 2 2.6 Protein analysis 2.6.
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Chapter 2 2.6.2- Native gel Reagent
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Chapter 2 - Diluted primary antibod
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Chapter 2 supernatant was decanted
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Chapter 2 2.6.6- Analytical ultrace
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Chapter 3 Background As discussed a
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Chapter 3 the -35 region of the trp
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Chapter 3 20 mins 40 mins 60 mins 8
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Chapter 3 (a) 60 mins 90 mins 120 m
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Chapter 3 For this experiment, MGM1
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Chapter 3 When IPTG was not include
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Chapter 3 The results from these co
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Chapter 3 3.2.1 The effect of chang
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Chapter 3 3.2.2 Using pRARE plasmid
Page 135 and 136: Chapter 3 3.3 Complementation and e
Page 137 and 138: Chapter 3 of MGM100. The use of Tab
Page 139 and 140: Chapter 3 3.3.1 Using pET plasmid i
Page 141 and 142: Chapter 3 1 2 3 4 5 6 7 8 95 kDa 72
Page 143 and 144: Chapter 3 it only digests the paren
Page 145 and 146: Chapter 3 72 kDa 1 2 3 4 5 55 kDa 1
Page 147 and 148: Chapter 3 30°C/26°C 37°C MGM100
Page 149 and 150: Chapter 3 Fig 3.12: Comparison of t
Page 151 and 152: Chapter 3 AI90/pACYC-Ptrc-GroESL-Sa
Page 153 and 154: Chapter 3 1 2 3 4 5 6 7 1000 bp 600
Page 155 and 156: Chapter 3 10 -1 10 -2 10 -3 10 -4 1
Page 157 and 158: Chapter 3 this observation. The res
Page 159 and 160: Chapter 3 expression. However, from
Page 161 and 162: Chapter 4 Construction and analysis
Page 163 and 164: Chapter 4 Background In the last ch
Page 165 and 166: Chapter 4 Experimental design To ma
Page 167 and 168: Chapter 4 Cpn60.2 7 8 9 Cpn60.1 1 2
Page 169 and 170: Chapter 4 The full list of all the
Page 171 and 172: Chapter 4 4.1 Imprecise domain swap
Page 173 and 174: Chapter 4 4.1.1 Chimera AEC This is
Page 175 and 176: Chapter 4 (a) Dilutions 10 -1 10 -2
Page 177 and 178: Chapter 4 of GroEL (fig 4.7). This
Page 179 and 180: Chapter 4 (a) Dilutions 10 -1 10 -2
Page 181 and 182: Chapter 4 4.1.5 Chimeras DHF & GEI
Page 183 and 184: Chapter 4 4.2 Precise domain swaps
Page 185: Chapter 4 intermediate domains were
Page 189 and 190: Chapter 4 Dilutions 10 -2 10 -3 10
Page 191 and 192: Chapter 4 is important to note that
Page 193 and 194: Chapter 4 Summary the complementati
Page 195 and 196: Chapter 4 The other aspect of this
Page 197 and 198: Chapter 4 caused due to lack of exp
Page 199 and 200: Chapter 4 structures were not expec
Page 201 and 202: Chapter 4 Name Construct Blue -Cpn6
Page 203 and 204: Chapter 5 Background As discussed i
Page 205 and 206: Chapter 5 macromolecules at equilib
Page 207 and 208: Chapter 5 5.1 Purification of JNL 5
Page 209 and 210: Chapter 5 (a) 1 2 3 4 5 6 7 8 9 10
Page 211 and 212: Chapter 5 (a) A B C D E (b) 1 2 3 4
Page 213 and 214: Chapter 5 5.2 AUC analysis of JNL U
Page 215 and 216: Chapter 5 (a) 1 JNL in Buffer 5 0.9
Page 217 and 218: Chapter 5 (a) (b) 100 mM P i 75 mM
Page 219 and 220: Chapter 5 In this experiment GroEL
Page 221 and 222: Chapter 5 1 2 3 Corrected values Av
Page 223 and 224: Chapter 5 expected, evidence of lar
Page 225 and 226: Chapter 5 In summary, based on the
Page 227 and 228: Chapter 6 In this chapter I will be
Page 229 and 230: Chapter 6 of Mscpn60.1 can indirect
Page 231 and 232: Chapter 6 can produce products with
Page 233 and 234: Chapter 6 As the group in Pittsburg
Page 235 and 236: Chapter 6 Fig 6.3: Masses of 4 day
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Chapter 6 than 7 days, so there is
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Chapter 6 are required to facilitat
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Chapter 6 Fig 6.4: Sequence alignme
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Chapter 6 To ensure that the lack o
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Chapter 6 6.2.3 Complementation ana
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Chapter 6 Discussion The double rin
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Chapter 6 Section 6.3: Testing the
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Chapter 6 Experimental design For t
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Chapter 6 EcoRV groEL (1.6kb) HindI
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Chapter 6 6.3.2 Complementation res
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Chapter 6 1 2 3 4 72 kDa 55 kDa Fig
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Chapter 6 10 -1 10 -2 10 -3 10 -4 1
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Chapter 6 Discussion The results th
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Summary The main aim of this study
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together, these results strongly su
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Basha, E., K. L. Friedrich & E. Vie
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Cehovin, A., A. R. Coates, Y. Hu, Y
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Diaz-Acosta, A., M. L. Sandoval, L.
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Fux, C. A., J. W. Costerton, P. S.
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Gupta, P., S. Mishra & T. K. Chaudh
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Hoffmann, A., F. Merz, A. Rutkowska
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Ito, K., Y. Akiyama, T. Yura & K. S
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Krzewska, J., G. Konopa & K. Libere
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Lin, Z., D. Madan & H. S. Rye, (200
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Mogk, A. & B. Bukau, (2004) Molecul
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Patzelt, H., G. Kramer, T. Rauch, H
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Rommelaere, H., M. Van Troys, Y. Ga
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Strickland, E., B. H. Qu, L. Millen
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Tyagi, N. K., W. A. Fenton & A. L.
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Yamamori, T. & T. Yura, (1982) Gene
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