Targeting Functional Centers of the Ribosome

<p>1. Introduction  </p><p>1.1 Ribosomal antibiotics: function, activity and selectivity           </p><p>1.2 Pleuromutilins: a class of PTC ribosomal antibiotics for clinical use   </p><p>1.3 Structural basis for cross-resistance between ribosomal PTC antibiotics          </p><p>1.4 Oligonucleotides as ribosomal inhibitors and as tools for structural and functional study       </p><p>1.5 Minimal ribosomal components with PTC structure and function       </p><p>2. Methods         </p><p>2.1 Structural study of pleuromutilin antibiotics               </p><p>2.1.1 Crystallization and data collection</p><p>2.1.2 Data processing, structure solution and refinement               </p><p>2.2 Comparative structural analysis to reveal the structural basis for cross-resistance between PTC antibiotics          </p><p>2.3 Antisense oligonucleotides for targeting functional ribosomal centers             </p><p>2.3.1 Database construction         </p><p>2.3.2 In-vitro transcription-translation system for ribosome activity assay            </p><p>2.3.3 Antisense oligonucleotides nomenclature   </p><p>2.4 Minimal ribosomal components with PTC structure and function      </p><p>2.4.1 In vitro RNA transcription </p><p>2.4.2 Study of dimerization tendency      </p><p>2.4.3 Electrophoresis Mobility Shift Assay (EMSA)           </p><p>2.4.4 Size Exclusion Chromatography (SEC) for the separation between dimer and monomer      </p><p>2.4.5 Radiolabeling of substrates for Peptidyl Transferase activity assay  </p><p>2.4.6 Assay for Peptidyl Transferase activity       </p><p>2.4.7 RNA two-dimensional structure prediction              </p><p>2.5 Numbering, Sequence Alignment, and Images.           </p><p>3. Results            </p><p>3.1 The structures of D50S/Pleuromutilins comlexes        </p><p>3.2 Structural basis for cross resistance between ribosomal PTC antibiotics          </p><p>3.3 Oligonucleotides as ribosomal inhibitors and as tools for structural and functional study       </p><p>3.3.1 Correlation between IC50 and various ODN parameters       </p><p>3.3.2. Effect of ODN length          </p><p>3.4 Minimal ribosomal components with PTC structure and function      </p><p>3.4.1 Construct design    </p><p>3.4.2 Study of dimerization tendency      </p><p>4. Discussion     </p><p>4.1 Pleuromutilins           </p><p>4.1.1 Induced-fit mechanism for pleuromutilin binding  </p><p>4.1.2 C14 extension is located in the PTC void.     </p><p>4.1.3 Pleuromutilins resistance   </p><p>4.1.4 Pleuromutilins selectivity acquired by remote interactions               </p><p>4.2 Structural basis for cross resistance between ribosomal PTC antibiotics          </p><p>4.2.1 Resistance to PTC antibiotics is frequently acquired by mutating remote nucleotides           </p><p>4.2.2 U2504 at the crossroad of remote mutations networks that hamper binding of PTC antibiotics.</p><p>4.2.3 Second layer nucleotides    </p><p>4.2.4 Third Layer Nucleotides    </p><p>4.2.5 Resistance to various PTC antibiotics mediated by the same nucleotides      </p><p>4.3 Oligonucleotides as ribosomal inhibitors and as tools for structural and functional studies    </p><p>4.4 Minimal ribosomal components with PTC structure and function     </p><p>5. References      </p><p>6. Figures, tables and plots          </p>