Structural Determination of ApoE(222-271) in lipid environment by NMR spectroscopy
| Autor: | Tzu-Yi Chang, 張子弈 |
|---|---|
| Rok vydání: | 2017 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 106 Human apolipoprotein E (apoE) is a polymorphic protein of 299 amino acids with a molecular weight of ~34 kDa. It consists of two independently structural domains and separated by a flexible hinge region. One of the structural domains is a 22 kDa N-terminal domain (residues 1-191) and the other one is 10 kDa C-terminal domain (residues 216-299). The N-terminal domain and C-terminal domain are responsible for LDL receptor and lipid binding activity, respectively. ApoE has three major isoforms, it plays a key role in lipid metabolism. They displayed significant differences in their biological functions. ApoE2 is associated with type III hyperlioproteinemia. ApoE4 has been thought to be a risk factor for Alzheimer’s disease and atherosclerosis. The interaction mechanism of apoE with lipid remains unclear. Our ultimate goal is to elucidate the mechanism of apoE and lipid interaction from the structural point of view. Previously, our laboratory reported the secondary structure of the C-terminal domain of apoE in the presence of lipid. However, the three-dimensional structure of apoE in the presence of lipid remains unknown. In this study, we perform structural determination on C-terminal fragment from residue 222 to 271 which is major lipid binding domain. The 3D structure of apoE(222-271) in the presence and absence of dihexanoyl phosphatidylcholine (DHPC) micelles was characterized by using nuclear magnetic resonance spectroscopy (NMR), cicular dicroismc (CD) and analytic ultracentrifugation (AUC) techniques. The structural conformation may help us gain insight into the mechanism of lipid clearance by apoE. Our data shows that monomeric form of apoE(222-271) in aqueous solution was obtained. In the presence of DHPC micelles, apoE(222-271) formed complex with DHPC micelles and reavled that lipid binding increased the α-helix content. ApoE(222-271) is getting low affinity with micelles after dilution below the CMC (critical micelle concentration), but still bind with micelles. And we use XPLOR to simulate the 3D structures of apoE(222-271) in the presence and absence of DHPC micelles. We find that both have two major structures. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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