| 摘要 | 第4-8页 |
| abstract | 第8-9页 |
| Chapter1.Introduction | 第12-28页 |
| 1.1 Current status of cytochrome P450s | 第12-14页 |
| 1.2 The emphasis on the human CYP4Z | 第14-16页 |
| 1.3 Different CYP-dependant test systems | 第16-19页 |
| 1.4 The“enzyme bag”method based on permeabilized S.pombe cells | 第19-21页 |
| 1.5 The advantages and applications of bioluminescent CYP assays | 第21-23页 |
| 1.6 Homology modelling and molecular docking | 第23-26页 |
| 1.7 The prodrug strategy for human CYP4Z | 第26-28页 |
| Chapter2.Materials and methods | 第28-38页 |
| 2.1 Chemicals and biological substance | 第28页 |
| 2.2 Instruments and consumables | 第28-29页 |
| 2.3 General techniques for fission yeast and Escherichia coli | 第29-30页 |
| 2.4 Expression plasmid construction | 第30页 |
| 2.5 Fission yeast strain construction | 第30-31页 |
| 2.6 Whole-cell biotransformation in shaking flasks | 第31-32页 |
| 2.7 Biotransformation with enzyme bags | 第32-34页 |
| 2.7.1 Preparation for enzyme bags | 第32页 |
| 2.7.2 Permeabilization of fission yeast cells | 第32页 |
| 2.7.3 Incubation procedures | 第32-34页 |
| 2.7.4 Stop and analysis | 第34页 |
| 2.8 Activity and inhibitor assay in human liver microsomes | 第34-35页 |
| 2.8.1 The effect of Triton X-100 on the on the CYP activity in HLMs | 第34-35页 |
| 2.8.2 The percentage inhibition of HLMs by six test compounds | 第35页 |
| 2.9 Statistical analysis | 第35页 |
| 2.10 Homology modeling of CYP4Z1 and molecular docking experiments | 第35-38页 |
| 2.10.1 Homology modeling of CYP4Z | 第35-36页 |
| 2.10.2 molecular docking experiments | 第36-38页 |
| Chapter3.Results | 第38-60页 |
| 3.1 Effect of Triton X-100 on the activity of CYP2C9 and CYP3A4 in Human liver microsomes | 第38-39页 |
| 3.2 Direct comparison of CYP3A4 activity in whole-cell biotransformation and enzyme bag-catalyzed biotransformation | 第39-40页 |
| 3.3 CYP2C9 activity in enzyme bags | 第40-42页 |
| 3.3.1 Time dependent assay of CYP2C9 activity by using enzyme bags | 第40-41页 |
| 3.3.2 Long time storage assay of CYP2C9 activity by using enzyme bags | 第41-42页 |
| 3.4 Identification of new CYP4Z1 probe substrates | 第42-44页 |
| 3.5 Identification of new CYP4Z1 inhibitors | 第44-48页 |
| 3.5.1 The inhibition assay of CYP4Z1 with six compounds | 第44-46页 |
| 3.5.2 Inhibitor specificity assessment in Human liver microsomes,CYP2C9-and CYP4Z1-expressing enzyme bags | 第46-47页 |
| 3.5.3 IC50 determination for inhibition of CYP4Z1 by1-benzylimidazole | 第47-48页 |
| 3.6 Homology modeling for CYP4Z | 第48-50页 |
| 3.7 Substrate and inhibitor docking experiments | 第50-58页 |
| 3.7.1 Substrates docking experiments | 第51-57页 |
| 3.7.2 Inhibitor docking experiments | 第57-58页 |
| 3.8 The effects of amino acid mutations for CYP4Z1 wild-type | 第58-60页 |
| CHAPTER4.DISCUSSIONS | 第60-70页 |
| 4.1 Influence of Triton X-100 on CYP activity | 第60-61页 |
| 4.2 Direct comparison of CYP3A4 activity in whole-cell biotransformation and enzyme bag-catalyzed biotransformation | 第61页 |
| 4.3 CYP2C9 activity in enzyme bags | 第61-62页 |
| 4.3.1 Time dependent assay of CYP2C9 activity by using enzyme bags | 第61-62页 |
| 4.3.2 Long time storage assay of CYP2C9 activity by using enzyme bags | 第62页 |
| 4.4 Identification of new CYP4Z1 probe substrates | 第62-63页 |
| 4.5 Homology modelling and substrates docking of CYP4Z | 第63-65页 |
| 4.6 Inhibitors identification and docking experiments of CYP4Z | 第65-67页 |
| 4.7 Mutagenesis assay | 第67-70页 |
| CHAPTER5.CONCLUSIONS AND PERSPECTIVES | 第70-72页 |
| REFERENCE | 第72-82页 |
| APPENDIX A | 第82-86页 |
| APPENDIX B | 第86-90页 |
| NOTES ON PUBLICATIONS AND PARTICIPATION IN SCIENTIFIC RESEARCH | 第90-92页 |
| ACKNOWLEDGEMENT | 第92页 |
