Purification and characterization of a 19 kDa zinc-binding protein in porcine brain.
by Wong Ping Shing. === Thesis (M.Phil.)--Chinese University of Hong Kong, 1995. === Includes bibliographical references (leaves 97-112). === ACKNOWLEDGMENTS --- p.i === ABSTRACT --- p.ii === ABBREVIATIONS --- p.viii === Chapter 1. --- INTRODUCTION --- p.1 === Chapter 1.1 --- General properties...
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| Format: | Others |
| Language: | English |
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Chinese University of Hong Kong
1995
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| Online Access: | http://library.cuhk.edu.hk/record=b5888510 http://repository.lib.cuhk.edu.hk/en/item/cuhk-320537 |
| Summary: | by Wong Ping Shing. === Thesis (M.Phil.)--Chinese University of Hong Kong, 1995. === Includes bibliographical references (leaves 97-112). === ACKNOWLEDGMENTS --- p.i === ABSTRACT --- p.ii === ABBREVIATIONS --- p.viii === Chapter 1. --- INTRODUCTION --- p.1 === Chapter 1.1 --- General properties of zinc === Chapter 1.1.1 --- Biochemistry of zinc --- p.2 === Chapter 1.1.2 --- Distribution of zinc in body --- p.3 === Chapter 1.1.3 --- Roles of zinc in protein function --- p.4 === Chapter 1.2 --- Zinc and zinc-binding proteins in brain === Chapter 1.2.1 --- Distribution of zinc in brain --- p.7 === Chapter 1.2.2 --- Metabolism of zinc in brain --- p.9 === Chapter 1.2.3 --- Compartments of zinc in brain --- p.10 === Chapter 1.2.4 --- Zinc-binding proteins in brain --- p.12 === Chapter 1.3 --- Pathological conditions of brain in relation to zinc --- p.15 === Chapter 1.4 --- Aim of the project --- p.20 === Chapter 2. --- MATERIALS AND METHODS --- p.22 === Chapter 2.1 --- Detection of zinc-binding proteins === Chapter 2.1.1 --- Sodium-Dodecyl Sulphate Polyacrylamide Gel Electrophoresis (SDS-PAGE) --- p.22 === Chapter 2.1.2 --- Electroblotting --- p.24 === Chapter 2.1.3 --- Radioactive zinc blotting --- p.25 === Chapter 2.1.4 --- Autoradiography --- p.25 === Chapter 2.2 --- Subcellular fractionation of porcine brain --- p.26 === Chapter 2.3 --- Purification and structural characterization of a 19 kDa zinc-binding protein === Chapter 2.3.1 --- Purification of a 19 kDa protein --- p.27 === Chapter 2.3.2 --- Sequencing of N-terminal blocked 19 kDa protein --- p.30 === Chapter 2.4 --- Characterization of the binding and biological properties of the 19 kDa zinc-binding protein === Chapter 2.4.1 --- Effect of divalent metal ions on zinc binding to the 19 kDa protein --- p.33 === Chapter 2.4.2 --- Effect of pH on the dissociation of radioactive zinc from the19 kDa protein --- p.34 === Chapter 2.4.3 --- Radioactive calcium blotting --- p.34 === Chapter 2.4.4 --- Interaction of radioactive zinc and radioactive calcium binding to the 19 kDa protein --- p.35 === Chapter 2.4.5 --- Calmodulin activity assay --- p.35 === Chapter 3. --- RESULTS === Chapter 3.1 --- Specificity of radioactive zinc-blot on zinc-binding protein detection --- p.38 === Chapter 3.2 --- Zinc-binding proteins in porcine brain --- p.38 === Chapter 3.3 --- Purification and identification of a cytosolic 19 kDa zinc- binding protein in porcine brain === Chapter 3.3.1 --- Zinc-dependent hydrophobic interaction chromatography --- p.44 === Chapter 3.3.2 --- N-terminal amino acid sequencing --- p.51 === Chapter 3.3.3 --- High pH native gel electrophoresis of 19 kDa protein --- p.51 === Chapter 3.4 --- The zinc and calcium binding properties of the 19 kDa protein === Chapter 3.4.1 --- Effect of pre-exposure to divalent cations on zinc binding --- p.54 === Chapter 3.4.2 --- Competition by divalent cations for zinc binding --- p.56 === Chapter 3.4.3 --- pH dependency of zinc dissociation --- p.56 === Chapter 3.4.4 --- Effect of zinc on radioactive calcium binding --- p.61 === Chapter 3.5 --- The biological activity of the 19 kDa protein === Chapter 3.5.1 --- Effect of the 19 kDa protein on the activity of calmodulin- dependent phosphodiesterase --- p.66 === Chapter 3.5.2 --- Effect of zinc on calmodulin-dependent phosphodiesterase activity --- p.69 === Chapter 3.5.4 --- "Effect of zinc on calcium-deficient, calmodulin-dependent phosphodiesterase activity" --- p.72 === Chapter 4. --- DISCUSSION === Chapter 4.1 --- Detection and Purification of zinc-binding proteins === Chapter 4.1.1 --- Strategy for the detection of zinc-binding proteins --- p.77 === Chapter 4.1.2 --- Purification of zinc-binding protein --- p.79 === Chapter 4.2 --- Amino acid sequencing of the 19 kDa protein --- p.82 === Chapter 4.3 --- Binding properties of the 19 kDa zinc-binding protein --- p.86 === Chapter 4.4 --- Effect of zinc and 19 kDa zinc-binding protein on calmodulin dependent phosphodiesterase --- p.92 === Chapter 4.5 --- Effect of zinc on the properties of calmodulin --- p.90 === Chapter 4.6 --- Significance of the ability of zinc to affect calmodulin activity --- p.94 === Chapter 5. --- CONCLUSION --- p.95 === Chapter 6. --- REFERENCES --- p.97 |
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