Genomics, proteomics, and vaccines

While the sequence of the human genome sequence has hit the headlines, extensive exploitation of this for practical applications is still to come. Genomic and post-genomic technologies applied to viral and bacterial pathogens, which are almost equally important from a scientific perspective, have the potential to be translated into useful products and processes much more rapidly. Genomics, Proteomics and Vaccines introduces the history of vaccinology and discusses how vaccines are expected to evolve in the future. It describes the relevant technologies, including genome sequencing and analysis, DNA microarrays, 2D electrophoresis and 2D chromatography, mass spectrometry and high-throughput protein expression and purification. The book also features examples of the exploitation of genomics and post-genomics in vaccine discovery, and contains useful descriptions of the biology and pathogenesis of clinically important bacterial pathogens. This book should be of interest to all those working in vaccine discovery and development in pharmaceutical and biotechnology companies as well as in academic institutionsContent: Chapter 1 Vaccination: Past, Present and Future (pages 1–22): Maria Lattanzi and Rino Rappuoli Chapter 2 Bioinformatics, DNA Microarrays and Proteomics in Vaccine Discovery: Competing or Complementary Technologies? (pages 23–41): Guido Grandi Chapter 3 Genome Sequencing and Analysis (pages 43–73): Herve Tettelin and Tamara Feldblyum Chapter 4 Understanding DNA Microarrays: Sources and Magnitudes of Variances in DNA Microarray Data Sets (pages 75–101): She?Pin Hung, Suman Sundaresh, Pierre F. Baldi and G. Wesley Hatfield Chapter 5 The Proteome, Anno Domini Two Zero Zero Three (pages 103–134): Pier Giorgio Righetti, Mahmoud Hamdan, Frederic Reymond and Joel S. Rossier Chapter 6 Mass Spectrometry in Proteomics (pages 135–169): Pierre?Alain Binz Chapter 7 High?Throughput Cloning, Expression and Purification Technologies (pages 171–182): Andreas Kreusch and Scott A. Lesley Chapter 8 Meningococcus B: From Genome to Vaccine (pages 183–204): Davide Serruto, Rino Rappuoli and Mariagrazia Pizza Chapter 9 Vaccines against Pathogenic Streptococci (pages 205–222): John L. Telford, Immaculada Margarit Y Ros, Domenico Maione, Vega Masignani, Herve Tettelin, Giuliano Bensi and Guido Grandi Chapter 10 Identification of the ‘Antigenome’ — A Novel Tool for Design and Development of Subunit Vaccines against Bacterial Pathogens (pages 223–243): Eszter Nagy, Tamas Henics, Alexander von Gabain and Andreas Meinke Chapter 11 Searching the Chlamydia Genomes for New Vaccine Candidates (pages 245–266): Giulio Ratti, Oretta Finco and Guido Grandi Chapter 12 Proteomics and Anti?Chlamydia Vaccine Discovery (pages 267–283): Gunna Christiansen, Svend Birkelund, Brian B. Vandahl and Allan C Shaw Chapter 13 Proteome Analysis of Outer Membrane and Extracellular Proteins from Pseudomonas aeruginosa for Vaccine Discovery (pages 285–304): Stuart J. Cordwell and Amanda S. Nouwens Figure 1......Page 19 Figure 3......Page 21 Figure 5......Page 22 Figure 7......Page 23 Figure 8......Page 24 Figure 9......Page 25 Figure 10......Page 26 1.1&X;Introduction......Page 28 1.2&X;Vaccination: the past......Page 29 1.3&X;Vaccination: the present......Page 31 1.3.1&Y;Conjugate vaccines......Page 32 1.3.2&Y;Recombinant DNA technology for subunit vaccines......Page 35 1.4.1&Y;‘Reverse vaccinology’......Page 37 1.4.2&Y;Improved delivery: mucosal vaccines......Page 38 1.5&X;Conclusion: the intangible value of vaccination......Page 39 mkr4......Page 40 mkr18......Page 41 mkr32......Page 42 mkr47......Page 43 mkr58......Page 44 2.1&X;Introduction......Page 48 2.2&X;From genome sequence to vaccine discovery......Page 50 Figure 1......Page 51 2.3&X;A case study: the anti-meningococcus B vaccine......Page 53 2.3.2&Y;The transcriptome analy?sis approach......Page 54 Table 1......Page 55 Figure 3......Page 58 Table 2......Page 59 Figure 2......Page 60 Figure 4......Page 61 2.5&X;Conclusions: a ‘nomics’ approach to vaccine discovery......Page 62 Figure 5......Page 64 mkr11......Page 65 mkr21......Page 66 3.1&X;Introduction......Page 68 3.2.1&Y;Library construction and template preparation......Page 69 Figure 1......Page 70 Figure 2......Page 72 3.2.2&Y;High-throughput sequencing......Page 73 Figure 3......Page 78 Equation 3......Page 79 3.2.4&Y;Genome finishing......Page 80 3.2.6&Y;Novel sequencing technologies......Page 82 3.3.1&Y;Annotation pipeline......Page 83 3.3.2&Y;Comparative genomics......Page 84 3.3.4&Y;Identification of vaccine candidates......Page 86 mkr2......Page 88 mkr17......Page 89 mkr31......Page 90 mkr44......Page 91 mkr48......Page 92 4.1&X;Introduction......Page 97 4.2.1&Y;&BI;In situ&N; synthesized oligonucleotide arrays......Page 98 4.2.3&Y;Filter-based DNA arrays......Page 99 Figure 1......Page 100 4.3&X;Data analy?sis meth?ods......Page 101 4.3.1&Y;Robust estimation of standard deviation with a small num?ber of replicates......Page 102 4.3.2&Y;Estimation of global false posi?tive levels......Page 103 Figure 3......Page 105 4.4.1&Y;Data sets......Page 106 Figure 4......Page 107 Figure 5......Page 108 4.4.2&Y;Data analy?sis......Page 109 4.4.4&Y;Correlations by pairs of experiments......Page 110 Table 1......Page 111 Table 2......Page 112 Table 3......Page 113 Table 4......Page 114 4.4.6&Y;Correlations be?tween GeneChip data processed with Affymetrix MAS 4.0, MAS 5.0 or dChip software......Page 115 Table 5......Page 117 Table 6......Page 118 Table 7......Page 119 mkr10......Page 122 mkr12......Page 123 5.1&X;Introduction......Page 124 Figure 1......Page 125 5.2&X;Some definitions......Page 126 5.3&X;What meth?ods exist to tackle the proteome complexity?......Page 128 5.3.1 Standard 2D map analy?sis......Page 129 5.4&X;Quantitative proteomics......Page 133 Figure 2......Page 134 Figure 3......Page 136 Figure 4......Page 137 5.5&X;Pre-fractionation in proteome analy?sis......Page 138 Figure 5......Page 139 Figure 6......Page 141 Figure 7......Page 143 5.7&X;Protein chip arrays......Page 144 Figure 8......Page 145 Figure 9......Page 146 5.8&X;Imaging mass spectrometry......Page 147 Figure 10......Page 148 mkr16......Page 149 mkr30......Page 150 mkr44......Page 151 mkr103......Page 155 6.1&X;Introduction......Page 156 6.2.1&Y;Ionization......Page 157 Figure 1......Page 158 Figure 2......Page 159 6.2.2&Y;Ion analy?sis......Page 160 Figure 3......Page 161 Figure 4......Page 162 Figure 5......Page 163 Figure 6......Page 165 6.2.3&Y;Tandem MS, MS/MS......Page 166 6.2.4&Y;Instrumentation......Page 167 6.3.1&Y;PMF and MS/MS......Page 168 Figure 7......Page 170 Figure 8......Page 172 Figure 9......Page 173 6.4&X;Proteomics workflows......Page 176 6.4.1&Y;The classical gel-based approach......Page 177 Figure 10......Page 178 6.4.2&Y;The molecular scanner approach......Page 179 Figure 11......Page 180 6.4.3&Y;The MuDPIT approach......Page 181 Figure 12......Page 182 6.4.4&Y;The ICAT approach......Page 183 Figure 13......Page 184 6.4.5&Y;The SELDI profiling approach......Page 185 mkr11......Page 186 mkr28......Page 187 mkr39......Page 188 7.1&X;Introduction......Page 191 7.2&X;Gene cloning......Page 192 7.3&X;Protein ex?pres?sion......Page 194 Figure 1......Page 195 Figure 2......Page 197 7.5&X;Validation of the pipeline and outlook......Page 198 Table 1......Page 199 mkr10......Page 200 mkr24......Page 201 mkr36......Page 202 8.1.1&Y;Microbiological features and pathogenesis......Page 204 Figure 1......Page 205 Figure 2......Page 206 8.1.2&Y;Prevention: state of the art on Neisseria vaccines......Page 207 8.2&X;Group B meningococcus as an example of reverse vaccinology......Page 209 8.2.2&Y;Experimental strategy......Page 210 Figure 3......Page 211 Figure 4......Page 212 8.2.3&Y;Functional characterization of novel candidates......Page 214 Figure 5......Page 216 8.3&X;Conclusions......Page 219 mkr17......Page 220 mkr35......Page 221 mkr54......Page 222 9.1&X;Introduction......Page 224 9.2&X;Comparative genomics of streptococci......Page 225 Figure 1......Page 226 9.3.1&Y;Rationale......Page 227 Figure 2......Page 228 9.3.3&Y;High-throughput cloning and ex?pres?sion......Page 229 Figure 3......Page 230 9.3.5&Y;Confirmation of protection......Page 231 Table 1......Page 232 9.3.6&Y;Strain variation in antigen sequences......Page 233 9.4&X;A vaccine against group A streptococcus......Page 234 9.5&X;Conclusions......Page 237 mkr1......Page 238 mkr13......Page 239 mkr26......Page 240 mkr27......Page 241 10.1&X;Introduction......Page 242 Table 1......Page 245 9.2&X;Small DNA insert libraries......Page 246 Figure 1......Page 248 10.3&X;Proper display platforms......Page 249 10.4&X;Selected human sera provide imprints of pathogen encounters......Page 250 Figure 2......Page 252 10.5&X;Cognate anti?bodies reveal the ‘antigenome’ of a pathogen......Page 253 9.6&X;How to retrieve the candidate antigens......Page 254 10.7&X;Summary and dis?cus?sion......Page 256 mkr1......Page 258 mkr16......Page 259 mkr31......Page 260 mkr43......Page 261 mkr50......Page 262 11.1&X;Old problems and new per?spec?tives for chlamydial vaccines......Page 263 11.2&X;Post-genomic ap?proaches......Page 268 11.3.1&Y;&BI;C. pneumoniae&N; vaccine candidates......Page 269 Figure 1......Page 270 Figure 2......Page 271 11.3.2&Y;&BI;C. trachomatis&N; vaccine candidates......Page 274 11.3.3&Y;&BI;In vivo&N; evaluations......Page 278 11.4&X;Concluding considerations......Page 280 mkr14......Page 281 mkr35......Page 282 mkr50......Page 283 mkr57......Page 284 12.1&X;Introduction......Page 285 12.2.1&Y;EBs proteins......Page 287 Figure 1......Page 289 12.2.2&Y;Comparison of EBs proteomes from dif?fer?ent serovars......Page 290 12.2.4&Y;IFN-......Page 291 12.2.5&Y;Induction of &BI;C. trachomatis&N; tryptophan synthase by IFN-......Page 292 12.2.6&Y;Protein compartmentalization......Page 293 12.2.7&Y;RB-specific proteins......Page 294 Figure 2......Page 295 12.3.2&Y;Small unrecognized ORFs......Page 296 12.4&X;Benefits that proteomics provide for vaccine develop?ment......Page 297 mkr8......Page 298 mkr20......Page 299 mkr34......Page 300 mkr44......Page 301 13.1&X;Introduction......Page 302 11.2&X;Membrane proteins in......Page 303 13.2.1&Y;Proteomics of &BI;P. aeruginosa&N; outer membrane proteins......Page 305 Figure 1......Page 306 Figure 2......Page 307 13.3&X;Extracellular proteins in......Page 309 13.3.1&Y;Proteomics of &BI;P. aeruginosa&N; extracellular proteins......Page 310 Figure 3......Page 312 Figure 4......Page 313 Figure 5......Page 314 13.5&X;Conclusions......Page 315 mkr12......Page 316 mkr28......Page 317 mkr42......Page 318 mkr53......Page 319

While the sequence of the human genome sequence has hit the headlines, extensive exploitation of this for practical applications is still to come. Genomic and post-genomic technologies applied to viral and bacterial pathogens, which are almost equally important from a scientific perspective, have the potential to be translated into useful products and processes much more rapidly. Genomics, Proteomics and Vaccines introduces the history of vaccinology and discusses how vaccines are expected to evolve in the future. It describes the relevant technologies, including genome sequencing and analysis, DNA microarrays, 2D electrophoresis and 2D chromatography, mass spectrometry and high-throughput protein expression and purification. The book also features examples of the exploitation of genomics and post-genomics in vaccine discovery, and contains useful descriptions of the biology and pathogenesis of clinically important bacterial pathogens. This book should be of interest to all those working in vaccine discovery and development in pharmaceutical and biotechnology companies as well as in academic institutions

The complete genome sequences of many bacteria are now available, and comparisons of pathogenic strains with non-pathogenic relatives are becoming more common. This will enable researchers to target vaccine development towards specific gene products that traditional approaches of vaccine research failed to discover. Today, there are several examples of vaccine candidate identification using genomics and proteomics, making "Genomics, Proteomics, and Vaccines's state-of-the-art picture of the field and summary of the possible strategies extremely useful