12.06 - Olson - Membranes

12.06 - Olson - Membranes - 1 Membranes 2 3 4 5 6 7...

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Unformatted text preview: 1 Membranes 2 3 4 5 6 7 Enzymatic Markers of Differrent Membranes •Membranes – Plasma – Endoplasmic Reticulum – Golgi Apparatus – Inner Mitochondrial •Enzyme – 5’ Nucleotidase – Adenylyl cyclase – Na + K + ATPase – Glucose-6-phosphatase – GlcNAc Transferase I – ATP Synthase Specific Activity 8 9 10 11 12 Artificial Membranes •Micelles •Bilayers •Liposomes 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 Liposomes 29 30 Biological Membranes •Lipids •Proteins •Carbohydrates 31 32 Lipids •Phospholipids •Glycolipids •Cholesterol 33 34 35 36 Analysis of Asymmetric Distribution of Phosphatidylethanolamine •Amino modifying reagents – Membrane impremeant 2,4,6- Trinitrobenzenesulfonic acid (TNBS) – Membrane permeant -1- fluoro-2,4,dinitrobenzene (FDNB) •Treat Cells with amino modifying reagents 37 •extract lipids •separation on thin layer chromatography •isolation of compound 38 •check absorbance at 410 nm and quantitate spectrophotometrically •HPLC Analysis - quantitation of peak •compare total PE (FDNB) vs. external PE (TNBS) 39 40 41 42 Proteins •Integral •Peripheral 43 44 45 Transmembrane Arrangement of Membrane Proteins • compounds that react with free amino groups on proteins • membrane permeant – Ethylacetimidate (EA) • membrane impremeant – isoethionylacetimidate (IEA) 46 Radiolabel EA and IEA with 14C; Treat cells with [14C]EA and [14C]IEA 47 EA vs. IEA 48 49 50 Hydropathy index •measurement of the overall hydrophobicity of a sequence of amino acids that is estimated by summing the...
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This note was uploaded on 04/18/2008 for the course BIOC 1010 taught by Professor Zhang during the Fall '07 term at New York Medical College.

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12.06 - Olson - Membranes - 1 Membranes 2 3 4 5 6 7...

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