| ABSTRACT | 第5-7页 |
| 摘要 | 第8-13页 |
| 1. Introduction | 第13-43页 |
| 1.1 Laccase structure and function | 第13-22页 |
| 1.1.1 Different bacterial laccases and their functions | 第13-16页 |
| 1.1.2 Structural characteristics of bacterial laccase | 第16-20页 |
| 1.1.3 Cellular localization and molecular characteristics ofbacterial laccase | 第20-22页 |
| 1.2 Catalytic characteristics of bacterial laccases | 第22-28页 |
| 1.2.1 Substrates of laccases | 第23-26页 |
| 1.2.2 Effect of inhibitors on laccase | 第26-27页 |
| 1.2.3 Laccase thermo-stability | 第27-28页 |
| 1.2.4 pH effect on laccase activity | 第28页 |
| 1.3 Bacterial laccase production and applications | 第28-38页 |
| 1.3.1 Environmental factors affecting bacterial laccase production | 第28-31页 |
| 1.3.2 Molecular biology tools for enhancing the production and enzymatic properties ofbacterial laccase | 第31-33页 |
| 1.3.3 Applications of bacterial laccases | 第33-38页 |
| 1.4 Laccase immobilization | 第38-41页 |
| 1.5 Aims and objectives | 第41-43页 |
| 2. CotA laccase expression in recombinant E. coli BL21(DE3) | 第43-63页 |
| 2.1 Introduction | 第43-44页 |
| 2.2 Materials and Methods | 第44-54页 |
| 2.2.1 Materials and strains | 第44页 |
| 2.2.2 CotA laccase gene isolation and insertion of TTA codon into gene sequence | 第44-49页 |
| 2.2.3 Cloning of CotA laccase gene with/without TTA codon insertion into pET30a andpT7-FLAG-MAT-TAG-1 expression vectors | 第49-51页 |
| 2.2.4 Expression and purification of pET30a-CotA and pT7-FLAG-MAT-TAG-1-CotAlaccases | 第51-53页 |
| 2.2.5 RNA Isolation and qRT-PCR quantification | 第53-54页 |
| 2.3 Results and discussion | 第54-62页 |
| 2.3.1 CotA laccase gene isolation and amplification | 第54-58页 |
| 2.3.2 Expression and purification of CotA laccase from both expression vectors | 第58-62页 |
| 2.4 Conclusions | 第62-63页 |
| 3. Development of efficient micro-aerobic cultivation strategy for enhancing CotAlaccase production | 第63-79页 |
| 3.1 Introduction | 第63-64页 |
| 3.2 Materials and Methods | 第64-68页 |
| 3.2.1 Materials | 第64页 |
| 3.2.2 Process optimization for CotA laccase expression | 第64-65页 |
| 3.2.3 Measurement of ROS production | 第65页 |
| 3.2.4 Analytical procedure | 第65-68页 |
| 3.2.5 Dye Decolorization test for CotA laccase | 第68页 |
| 3.3 Results and discussion | 第68-78页 |
| 3.3.1 Process optimization for CotA laccase production under aerobic cultivation | 第68-71页 |
| 3.3.2 CotA laccase production under the micro-aerobic condition | 第71-75页 |
| 3.3.3 Dye decolorization using CotA laccase | 第75-78页 |
| 3.4 Conclusions | 第78-79页 |
| 4. CotA laccase immobilized on functionalized magnetic graphene oxide nano-sheets forefficient bio-catalysis | 第79-104页 |
| 4.1 Introduction | 第79-80页 |
| 4.2 Experimental section | 第80-84页 |
| 4.2.1 Materials | 第80页 |
| 4.2.2 Preparation of magnetic graphene oxide | 第80-81页 |
| 4.2.3 Preparation of MGO-NTA-Cu~(2+) nano-sheets | 第81页 |
| 4.2.4 Characterization of the synthesized nano-sheets | 第81-82页 |
| 4.2.5 CotA laccase immobilization | 第82页 |
| 4.2.6 Activity assay of free and immobilized CotA laccase | 第82-83页 |
| 4.2.7 Determination of immobilized CotA laccase catalytic characteristics | 第83页 |
| 4.2.8 Decolorization using the immobilized CotA laccase | 第83-84页 |
| 4.3 Results and Discussion | 第84-103页 |
| 4.3.1 Fabrication and characterization of MGO-NTA-Cu~(2+) nano-sheets andMGO-NTA-Cu-CotA | 第84-92页 |
| 4.3.2 CotA laccase immobilization | 第92-94页 |
| 4.3.3 Catalytic characteristics of immobilized CotA laccase | 第94-97页 |
| 4.3.4 Kinetic studies of free and immobilized CotA laccase | 第97-99页 |
| 4.3.5 Decolorization using MGO-NTA-Cu-CotA laccase | 第99-103页 |
| 4.4 Conclusions | 第103-104页 |
| 5. Conclusions and future perspectives | 第104-106页 |
| 5.1 Conclusions | 第104-105页 |
| 5.2 Novelty | 第105页 |
| 5.3 Future perspectives | 第105-106页 |
| List of abbreviations | 第106-108页 |
| References | 第108-132页 |
| Curriculum vitae | 第132-134页 |
| Acknowledgements | 第134页 |
