Fluorescence Studies on Biological Membranes

As stated by its first editor, Dr. D. B. Roodyn, the primary goal of the series Subcellular Biochemistry is to achieve an integrated view of the cell by bringing together results from a wide range of different techniques and disciplines. This volume deals with the applications of fluorescence spectroscopy to membrane research. It seeks to present complementary biochemical and bio­ physical data on both the structure and the dynamics of biological membranes. Biophysics and biochemistry are improving more and more in their ability to study biomembranes, overlapping somewhat in this area and explaining the functioning of the whole cell in terms of the properties of its individual com­ ponents. Therefore, we have brought together an international group of experts in order to report on and review advances in fluorescence studies on biological membranes, thereby highlighting subcellular aspects. The first chapters present a critical evaluation of the current applications of dynamic and steady-state fluorescence techniques. Subsequent chapters dis­ cuss more specific applications in cells, biological membranes, and their con­ stituents (lipids, proteins).

Inhalt

1 Biomembrane Structure and Dynamics Viewed by Fluorescence.- 1. Introduction to Fluorescence.- 2. Dynamics and Structure of Membranes.- 2.1. Lateral and Rotational Diffusion.- 2.2. Orientational Order and Packing.- 2.3. Asymmetry.- 2.4. Lipid Domains.- 3. Fluorescence Techniques and What They Make Visible.- 3.1. Fluorescence Depolarization.- 3.2. Quenching.- 3.3. Fluorescence Energy Transfer.- 3.4. Fluorescence Recovery after Photobleaching (FRAP).- 3.5. Excimer Fluorescence.- 4. Summary and Conclusions.- 5. References.- 2 Dynamic Structure of Membranes and Subcellular Components Revealed by Optical Anisotropy Decay Methods.- 1. Introduction.- 2. Optical Anisotropy Decay.- 2.1. Principle of Optical Anisotropy Decay Method.- 2.2. Experimental Techniques.- 2.3. Information Contained in an Anisotropy Decay.- 2.4. Optical Anisotropy Decay as a Tool in Bioscience.- 3. Examples of Application.- 3.1. Dynamic Structure of Membranes Probed by Diphenylhexatriene.- 3.2. Protein Rotations in Membrane and on Membrane Surface.- 3.3. Internal Motion of DNA.- 3.4. Internal Motion of Actin Filament.- 3.5. Dynamic Structure of Myosin Filament.- 4. Concluding Remarks.- 5. References.- 3 Principles of Frequency-Domain Fluorescence Spectroscopy and Applications to Cell Membranes.- 1. Introduction.- 2. Comparison of Time- and Frequency-Domain Measurements.- 2.1. A First-Generation Frequency-Domain Fluorometer.- 2.2. Resolution of a Two-Component Mixture.- 3. Theory of Frequency-Domain Fluorometry.- 3.1. Decays of Fluorescence Intensity.- 3.2. Decays of Fluorescence Anisotropy.- 4. Intensity Decays of DPH-Labeled Membranes.- 5. Anisotropy Decays of Labeled Membranes.- 5.1. Hindered Rotations of Diphenylhexatriene.- 5.2. Anisotropic Rotations of Perylene.- 6. Time-Resolved Emission Spectra.- 6.1. Calculation of Time-Resolved Emission Spectra.- 6.2. Time-Resolved Emission Centers of Gravity and Spectral Half-Widths.- 6.3. Time-Resolved Spectral Data for Patman-Labeled Membranes.- 7. Energy Transfer in Membranes.- 7.1. Distribution of Distances in a Covalently Linked Donor-Acceptor Pair.- 8. A 2-GHz Frequency-Domain Fluorometer.- 8.1. Picosecond Resolution of Tyrosine Intensity and Anisotropy Decays.- 8.2. Measurement of a 8-psec Correlation Time.- 9. Future Developments.- 10. Summary.- 11. References.- 4 Time-Resolved Fluorescence Depolarization Techniques in Model Membrane Systems: Effect of Sterols and Unsaturations.- 1. Introduction.- 2. Intrinsic Motional Properties of Some Widely Used Fluorescent Probes.- 2.1. Motional Characteristics.- 2.2. Excited-State Characteristics.- 3. Sterol-Phospholipid Interactions in Model Membranes.- 3.1. Cholesterol-Phospholipid Interactions: Lecithin as Bilayer Matrix.- 3.2. Cholesterol-Phospholipid Interactions: Phospholipids Other Than Lecithin as Bilayer Matrix.- 3.3. Cholesterol Chemical Modification: Effect on Phospholipid Fatty Acyl Chains Order and Dynamics.- 4. Concluding Remarks.- 5. References.- 5 Fluorescence Polarization to Evaluate the Fluidity of Natural and Reconstituted Membranes.- 1. Introduction.- 1.1. Aims and Scope of This Chapter.- 1.2. Mechanism of Action and Biological Significance of Fluorescence Polarization Measurements of Membrane Fluidity.- 2. Methodology.- 2.1. Theory of Fluorescence Polarization for Ion-Membrane Measurements.- 2.2. Probe-Membrane Interactions.- 2.3. Probe-Ion Interactions.- 3. Current Advancements in the Measurement of Ion-Membrane Interactions Using Fluorescence Polarization.- 3.1. Natural Membranes.- 3.2. Reconstituted Membranes.- 4. Critical Evaluation of the Significance of Ion-Membrane Measurements.- 4.1. Advantages of Fluorescence Polarization for Evaluation of Ion-Membrane Interactions.- 4.2. Limitations of Fluorescence Polarization for Measurement of Ion-Membrane Interactions.- 4.3. Substantiation of the Fluorescence Polarization Measurements of Ion-Membrane Interactions.- 5. Concluding Remarks.- 6. References.- 6 Fluidity of Thyroid Plasma Membranes.- 1. Introduction.- 2. Thyroid Plasma Membranes.- 2.1. Enriched Plasma Membrane Fractions.- 2.2. Chemical Characterization of Purified Plasma Membranes.- 2.3. Enzymic Characterization of Purified Plasma Membranes.- 2.4. Subfractionation of Thyroidal Plasma Membranes.- 2.5. Characterization of Thyroid Plasma Membrane Subfractions.- 3. Fluidity of Thyroid Plasma Membranes.- 3.1. Fluidity Measurements.- 3.2. Fluidity of Thyroid Subcellular Fractions.- 3.3. Fluidity of a P2 Fraction in Reconstituted Thyroid Plasma Membranes.- 3.4. Fluidity Characteristics of Plasma Membrane Subfractions.- 4. Modulation of the Adenylate Cyclase Activity by Manipulating the Plasma Membrane Composition.- 4.1. Incorporation of Phospholipids.- 4.2. Incorporation of Gangliosides.- 4.3. Incorporation of Dolichol and Dolichyl Derivatives.- 4.4. Addition of Membrane-Perturbing Drugs.- 5. Involvement of Membrane Fluidity on Human Normal and Pathological Thyroid Glands.- 6. References.- 7 Spectroscopic Analysis of the Structure of Bacteriorhodopsin.- 1. Introduction.- 2. Principle of the Fluorescence Energy Transfer Technique.- 3. Three-Dimensional Disposition of the Retinal Chromophore in the Purple Membrane.- 3.1. In-Plane Location.- 3.2. Transmembrane Location.- 3.3. Orientation of the Molecular Plane.- 4. In-Plane Location of NBD (7-Chloro-4-Nitrobenzo-2-Oxa-l,3-Diazole) Bound to Lys-41 in the Purple Membrane.- 5. Conformational Prediction of Bacteriorhodopsin Molecule.- 6. References.- 8 Structure and Dynamics of the Liver Microsomal Monoxygenase System.- 1. Introduction.- 1.1. General Structure of Biological Membranes.- 1.2. Peroxidation of Membrane Lipids.- 1.3. Microsomal Monoxygenase.- 2. Membrane Dynamics and Order Studied by Fluorescence.- 2.1. Biophysical Consequences of Lipid Peroxidation.- 2.2. Mobility of Membrane-Bound Cytochrome P-450.- 2.3. Rotational Mobility of Cytochrome P-450 in Peroxidized Rat Liver Microsomes.- 2.4. Structure of Free and Membrane-Bound Cytochrome P-450.- 2.5. Structure of NADPH-Cytochrome P-450 Reductase.- 2.6. Interaction of Cytochrome P-450 and Its Reductase in Membranes.- 2.7. Lipid-Protein Interactions Studied by DPH Fluorescence Anisotropy.- 3. References.- 9 Fluorescence Studies on Prokaryotic Membranes.- 1. Introduction.- 2. Fluorescent Probes.- 3. Structural Aspects of Bacterial Membranes.- 3.1. Outer Membrane of Gram-Negative Bacteria.- 3.2. Molecular Interactions.- 3.3. Phase Transitions and Homeoviscous Adaptation.- 3.4. Effects of Alcohols.- 3.5. Permeability of the Outer Membrane to Hydrophobic Substances.- 3.6. Membrane-Potential-Related Permeability Changes.- 3.7. Factors Increasing Cell Resistance and Membrane Stability.- 4. Periplasm.- 5. Incorporation of Exogenous Lipids into Prokaryotic Membranes.- 5.1. Gram-Negative Bacteria.- 5.2. Other Bacte…