유전자 정보를 바탕으로 한 Synechocystis sp. PCC6803의 대사재설계

Title
유전자 정보를 바탕으로 한 Synechocystis sp. PCC6803의 대사재설계
Authors
홍성주
Keywords
유전자정보를바탕으로한synechocystissppcc6803의대사재설계
Issue Date
2011
Publisher
인하대학교
Abstract
Cyanobacteria are the photosynthetic prokaryotes that contain photosystem I and II like eukaryotic microalgae and higher plants. They are also able to produce light-harvesting macromolecules such as chlorophyll, carotenoid and phycobiliproteins. Among cyanobacteria, Synechocystis sp. PCC 6803 is the first cyanobacteria to be fully sequenced. To establish in silico model of this microorganism based on whole genome sequence, the first step is to identify all the open reading frames and to determine their biochemical reactions with gene-protein-reaction association information. After all the network gaps are filled by biochemical or physiological evidences, the accuracy of metabolic network can be analyzed by mathematical methods such as flux-balance analysis. Prior to whole genome metabolic reconstruction, the central metabolism was built to confirm energy production and synthesize biomass precursors. ATP synthesis in photosynthesis was accomplished by pumping H+ ion from lumen to stromal side of the membrane in thylakoid membrane. Based on the above information, the central metabolism of Synechocystis PCC6803 was reconstructed using 78 genes and 56 biochemical reactions. The model was validated by simulation with respect to optimal growth yield. The simulation results correspond to the experimental data under various medium compositions and flux distribution in the network reactions. The 2nd central metabolism of Synechocystis sp. PCC6803 was remodelled with 82 reactions and 147 genes. Photosynthesis and respiration were expressed in more detail. As a result of simulation, under phototrophic culture Synechocystis utilized two biosynthetic pathways to generate biomass precursors instead of production of energy. However, in the case of heterotrophic culture, they produced energy using glyoxylate cycle. Based on this central metabolism, in silico model (iSJ716) was reconstructed with 716 biochemical reactions and 719 metabolites. It contains the characteristics of plan
Description
1.INTRODUCTION 1 2.LITERATURE SURVEY 5 2.1. Photosynthesis 5 2.1.1.Light-Dependent Reaction 7 2.1.2.Dark-Dependent Reaction 11 2.2.Cyanobacteria 14 2.2.1.Synechocystis sp. PCC6803 as the Model Microorganism 20 2.3. OMICs for Synechocystis sp. PCC6803 24 2.3.1.Genomics 24 2.3.2.Proteomics 26 2.3.3.Metabolomics 30 2.3.4.Transcriptomics 33 2.4.Systems Biology of Synechocystis sp. PCC6803 36 2.4.1.Bioinformatics 36 2.4.2.Systems biology 39 2.5.Aim of the Work 42 3.MATERIALS AND METHODS 44 3.1.Metabolic Reconstruction of Central Metabolism 44 3.1.1.Establishment of Metabolic Network 44 3.1.2.Mathematical Simulation 46 3.2.Whole Genome Metabolic Reconstruction 48 3.2.1.Construction of Metabolism 48 3.2.2.Determination of Biomass Object Function 49 3.3.Culture Conditions and Analytical Methods 51 3.3.1.Strain 51 3.3.2.Culture Media 51 3.3.3.Bubble Column Photobioreactor 52 3.3.4.Analytical Methods 52 3.4.Gene Deletion Analysis 54 3.4.1.Construction of Knockout Strains 54 3.4.2.Preparation of Insert DNA by Megaprimer PCR 54 3.4.3.Southern Blot Analysis 56 4.RESULTS AND DISCISSION 57 4.1.Evaluation of Central Metabolism under Various Trophic Conditions 57 4.1.1.Introduction 57 4.1.2.Photosynthetic Metabolism 59 4.1.3.Metabolic Reconstruction of Central Metabolism 63 4.1.4.Flux Analysis of Central Metabolism 66 4.1.5.Simulation of the Model 71 4.1.6.Conclusion 77 4.2.Analysis of Central Metabolism under Different Growth Modes 79 4.2.1.Introduction 79 4.2.2.Remodelling of the Central Metabolism in Synechocystis sp. PCC6803 82 4.2.3.The Role of Citric Acid Cycle for Different Growth Modes 85 4.2.4.Phenotype Phase Plane Analysis 93 4.2.5.Comparative Study among Citric Acid Cycles of Synechocystis and Other Bacteria 97 4.2.6.Conclusion 100 4.3.Metabolic Network on Whole Genome Sequence 102 4.3.1.Introduction 102 4.3.2.Reconstruction of the iSJ716 Model 104 4.3.3.Comparison of iSJ716 with Previously
URI
http://dspace.inha.ac.kr/handle/10505/22532
Appears in Collections:
College of Engineering(공과대학) > Biotechnology (생명공학) > Theses(생명공학 석박사 학위논문)
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