1st Edition

Membrane Analysis

ISBN 9781872748887
Published July 14, 1997 by Garland Science
224 Pages

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Book Description

This book provides a comprehensive review of laboratory methods for membrane study, with an emphasis on isolating membranes, analysing their composition and architecture, and investigating membrane function. Membrane Analysis describes the principles unde

Table of Contents

Abbreviations -- Preface -- 1. General membrane composition and structure -- Lipids -- General lipid composition -- Phospholipids -- Isoprenoids -- Lipid architecture -- Proteins -- Peripheral proteins -- Integral proteins -- Lipid-anchored proteins -- Protein architecture in the plane of the membrane -- Carbohydrates -- Glycolipids -- Glycoproteins -- Proteoglycans -- 2. The membranes of prokaryotes and eukaryotes -- General features -- Membranes of prokaryotes -- Membranes of eukaryotes -- The surface (plasma) membrane -- Plasma membrane domains -- The cytoskeleton and glycocalyx -- Mitochondria and chloroplasts -- Peroxisomes (microbodies) -- Lysosomes -- Nuclei -- Endocytic compartment -- The secretory compartment -- References -- 3. Preparation of subcellular membranes -- Homogenization -- Liquid shear -- Gaseous shear -- Mechanical shear -- Other methods -- Homogenization of soft mammalian tissue -- Homogenization of tough tissues -- Homogenization of cultured mammalian cells -- Plant tissue -- Yeast -- Bacteria -- Fractionation of subcellular organelles -- Differential centrifugation -- Density gradient centrifugation -- Density perturbation -- Fractionation by immunoaffinity -- Fractionation by continuous flow electrophoresis -- Fractionation in aqueous two-phase systems -- Identification of fractionated membranes -- Morphological -- Chemical -- Enzyme markers -- Immunological markers -- Expression of data -- Some specimen separations of organelles and membranes -- Nuclei -- Golgi membranes -- Mitochondria -- Resolving a light mitochondrial fraction -- Plasma membrane -- Membrane vesicles -- References -- 4. Molecular composition -- Chemical analysis of proteins -- Quantitative assay -- Separation of membrane proteins by sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE) -- Western blotting (immunoblotting) -- Resolution of proteins by isoelectric focusing and twodimensional (2D) electrophoresis -- Chemical analysis of lipids -- Lipid extraction -- Analysis of phospholipids -- Separation and analysis of phospholipids by HPLC -- Analysis of phospholipid fatty acid -- Determination of cholesterol -- Separation and analysis of glycolipids -- Glycoprotein oligosaccharide analysis -- Glycoprotein analysis -- Preparation of a crude glycoprotein fraction -- Glycoprotein fractionation -- Chemical assays for carbohydrates -- Isolation of the oligosaccharide chains -- Oligosaccharide sequence analysis -- References -- 5. Investigating the topology of membrane proteins -- Demonstration of the orientation of membrane vesicles -- Identification and separation of peripheral and integral proteins -- Triton X114 as a reagent for separation of integral and peripheral proteins -- Identification of proteins anchored by phosphatidylinositol -- Identification of lipid-modified proteins -- Probing the topography of integral membrane proteins -- General experimental criteria -- Probes for integral membrane proteins -- Hydropathy plots for prediction of membrane protein topography -- Structural analysis of membrane proteins -- References -- 6. Investigation of membrane lipids -- Determination of the organization of membrane phospholipids -- Model membranes -- Multilamellar vesicles (MLVs) -- Small unilamellar vesicles (SUVs) -- Large unilamellar vesicles (LUVs) -- Black lipid membranes -- Reconstituted membranes -- Determination of the transverse distribution of membrane phospholipids -- Chemical probes -- Phospholipases -- Phospholipid transfer proteins -- Assay based on prothrombinase -- Determination of the transverse distribution of membrane cholesterol -- Studies of lipid transport -- Investigation of the transmembrane movement of phospholipids -- Transport of phospholipids and cholesterol between intracellular membranes -- Genetic approach to investigate membrane phospholipids -- References -- 7. Protein targetting and membrane biogenesis -- Genetic approaches to studies of protein targetting -- Biochemical approaches to studies of protein targetting -- Steps in protein targetting -- Nuclear targetting -- Targetting to peroxisomes -- Targetting to mitochondria -- Targetting to the ER -- Transport across the ER membrane -- Retention in the ER lumen -- Transfer to the Golgi apparatus -- Targetting to lysosomes -- Transit of mannose-6-phosphatase receptors to and from the plasma membrane and Golgi membranes -- Establishing the topography and location of the membrane proteins of the ER, Golgi and plasma membrane -- Covalently bound lipids in protein targetting -- Targetting in prokaryotes -- Overview -- References -- 8. Endocytosis and vesicular trafficking -- Receptors and endocytosis -- Receptor-mediated endocytosis -- Membrane receptors -- Investigation of events in endocytosis -- Electron microscopy -- Biochemical studies -- Isolation of transport vesicles and investigation of the factors involved in vesicular transport -- Reconstitution of vesicular transport -- Semi-intact cell systems -- Transfer vesicles -- Transfer from the plasma membrane to the endosomes -- Transfer from the Golgi to lysosomes -- Golgi to endoplasmic reticulum (retrograde transport) -- Endoplasmic reticulum to Golgi (anterograde transport) -- Recognition in vesicular transfer -- References -- Index.

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John Graham Research Consultancy, Peter Jost Enterprise Centre, Liverpool John Moores University, Byrom Street, Liverpool L3 3AF, UK. The University of Sheffield, Department of Molecular Biology and Biotechnology, P.O. Box 594, Firth Court, Western Bank, Sheffield Sl0 2TN, UK.


"This book is primarily aimed at the newcomer to this field but the more experienced worker should find that it serves as a useful update." Nature Product Review; ; "Serving as a comprehensive review of laboratory protocols for membrane analysis, this pu