Pumps, channels, and transporters : methods of functional analysis /: methods of functional analysis. (2015)

• Record Type: Book
• Further Information: Note: Edited by Ronald J. Clarke, Mohammed A.A. Khalid.
• Editors: Clarke, Ronald J (Ronald James); Khalid, Mohammed A. A (A. Ali)
• Contents: Preface xv List of Contributors xix 1 Introduction 1; Mohammed A. A. Khalid and Ronald J. Clarke 1.1 History 1 1.2 Energetics of Transport 6 1.3 Mechanistic Considerations 7 1.4 Ion Channels 8 1.4.1 Voltage-Gated 8 1.4.2 Ligand-Gated 9 1.4.3 Mechanosensitive 9 1.4.4 Light-Gated 9 1.5 Ion Pumps 10 1.5.1 ATP-Activated 10 1.5.2 Light-Activated 11 1.5.3 Redox-Linked 12 1.6 Transporters 13 1.6.1 Symporters and Antiporters 13 1.6.2 Na+-Linked and H+-Linked 14 1.7 Diseases of Ion Channels, Pumps, and Transporters 15 1.7.1 Channelopathies 15 1.7.2 Pump Dysfunction 17 1.7.3 Transporter Dysfunction 18 1.8 Conclusion 18 References 19 2 Study of Ion Pump Activity Using Black Lipid Membranes 23; Hans - J ü rgen Apell and Valerij S. Sokolov 2.1 Introduction 23 2.2 Formation of Black Lipid Membranes 24 2.3 Reconstitution in Black Lipid Membranes 25 2.3.1 Reconstitution of Na+, K+-ATPase in Black Lipid Membranes 25 2.3.2 Recording Transient Currents with Membrane Fragments Adsorbed to a Black Lipid Membrane 26 2.4 The Principles of Capacitive Coupling 28 2.4.1 Dielectric Coefficients 29 2.5 The Gated-Channel Concept 31 2.6 Relaxation Techniques 34 2.6.1 Concentration-Jump Methods 34 2.6.2 Charge-Pulse Method 39 2.7 Admittance Measurements 39 2.8 The Investigation of Cytoplasmic and Extracellular Ion Access Channels in the Na+, K+-ATPase 42 2.9 Conclusions 43 References 45 3 Analyzing Ion Permeation in Channels and Pumps Using Patch - Clamp Recording 51; Andrew J. Moorhouse, Trevor M. Lewis, and Peter H. Barry 3.1 Introduction 51 3.2 Description of the Patch-Clamp Technique 52 3.2.1 Development of Whole-Cell Dialysis with Voltage-Clamp 52 3.3 Patch-Clamp Measurement and Analysis of Single Channel Conductance 54 3.3.1 Conductance and Ohm’s Law 54 3.3.2 Conductance of Channels versus Pumps 56 3.3.3 Fluctuation Analysis 57 3.3.4 Single Channel Recordings 61 3.4 Determining Ion Selectivity and Relative Permeation in Whole-Cell Recordings 67 3.4.1 Dilution Potential Measurements 67 3.4.2 Bi-Ionic Potential Measurements 69 3.4.3 Voltage and Solution Control in Whole-Cell Patch-Clamp Recordings 70 3.4.4 Ion Shift Effects During Whole-Cell Patch-Clamp Experiments 71 3.4.5 Liquid Junction Potential Corrections 72 3.5 Influence of Voltage Corrections in Quantifying Ion Selectivity in Channels 74 3.5.1 Analysis of Counterion Permeation in Glycine Receptor Channels 74 3.5.2 Analysis of Anion-Cation Permeability in Cation-Selective Mutant Glycine Receptor Channels 75 3.6 Ion Permeation Pathways through Channels and Pumps 76 3.6.1 The Ion Permeation Pathway in Pentameric Ligand-Gated Ion Channels 76 3.6.1.1 Extracellular and Intracellular Components of the Permeation Pathway 78 3.6.1.2 The TM2 Pore is the Primary Ion Selectivity Filter 79 3.6.2 Ion Permeation Pathways in Pumps Identified Using Patch-Clamp 80 3.6.2.1 Palytoxin Uncouples the Occluded Gates of the Na+, K+-ATPase 81 3.7 Conclusions 82 References 83 4 Probing Conformational Transitions of Membrane Proteins with Voltage Clamp Fluorometry (VCF) 89; Thomas Friedrich 4.1 Introduction 89 4.2 Description of The Vcf Technique 90 4.2.1 Generation of Single-Cysteine Reporter Constructs, Expression in Xenopus laevis Oocytes, Site-Directed Fluorescence Labeling 90 4.2.2 VCF Instrumentation 91 4.2.3 Technical Precautions and Controls 93 4.3 Perspectives from Early Measurements on Voltage-Gated K+ Channels 95 4.3.1 Early Results Obtained with VCF on Voltage-Gated K+ Channels 95 4.3.2 Probing the Environmental Changes: Fluorescence Spectra, Anisotropy, and the Effects of Quenchers 98 4.4 Vcf Applied to P-Type Atpases 100 4.4.1 Structural and Functional Aspects of Na+, K+- and H+, K+-ATPase 100 4.4.2 The N790C Sensor Construct of Sheep Na+, K+-ATPase α1-Subunit 102 4.4.2.1 Probing Voltage-Dependent Conformational Changes of Na+, K+-ATPase 103 4.4.2.2 The Influence of Intracellular Na+ Concentrations 107 4.4.3 The Rat Gastric H+, K+-ATPase S806C Sensor Construct 108 4.4.3.1 Voltage-Dependent Conformational Shifts of the H+, K+-ATPase Sensor Construct S806C During the H+ Transport Branch 109 4.4.3.2 An Intra- or Extracellular Access Channel of the Proton Pump? 110 4.4.3.3 Effects of Extracellular Ligands: K+ and Na+ 111 4.4.4 Probing Intramolecular Distances by Double Labeling and FRET 113 4.5 Conclusions and Perspectives 116 References 117 5 Patch Clamp Analysis of Transporters via Pre - Steady - State Kinetic Methods 121; Christof Grewer 5.1 Introduction 121 5.2 Patch Clamp Analysis of Secondary-Active Transporter Function 122 5.2.1 Patch Clamp Methods 122 5.2.2 Whole-Cell Recording 124 5.2.3 Recording from Excised Patches 124 5.3 Perturbation Methods 125 5.3.1 Concentration Jumps 126 5.3.2 Voltage Jumps 129 5.4 Evaluation and Interpretation of Pre-Steady-State Kinetic Data 130 5.4.1 Integrating Rate Equations that Describe Mechanistic Transport Models 131 5.4.2 Assigning Kinetic Components to Elementary processes in the Transport Cycle 131 5.5 Mechanistic Insight into Transporter Function 133 5.5.1 Sequential Binding Mechanism 133 5.5.2 Electrostatics 134 5.5.3 Structure-Function Analysis 134 5.6 Case Studies 136 5.6.1 Glutamate Transporter Mechanism 136 5.6.2 Electrogenic Charge Movements Associated with the Electroneutral Amino Acid Exchanger ASCT2 137 5.7 Conclusions 139 References 139 6 Recording of Pump and Transporter Activity Using Solid - Supported Membranes (SSM - Based Electrophysiology) 147; Francesco Tadini - Buoninsegni and Klaus Fendler 6.1 Introduction 147 6.2 The Instrument 148 6.2.1 Rapid Solution Exchange Cuvette 149 6.2.2 Setup and Flow Protocols 150 6.2.3 Protein Preparations 151 6.2.4 Commercial Instruments 152 6.3 Measurement Procedures, Data Analysis, and Interpretation 152 6.3.1 Current Measurement, Signal Analysis, and Reconstruction of Pump Currents 152 6.3.2 Voltage Measurement 156 6.3.3 Solution Exchange Artifacts 157 6.4 P-Type Atp ases Investigated by Ssm-Based Electrophysiology 159 6.4.1 Sarcoplasmic Reticulum Ca2+-ATPase 159 6.4.2 Human Cu+-ATPases ATP7A and ATP7B 163 6.5 Secondary Active Transporters 165 6.5.1 Antiport: Assessing the Forward and Reverse Modes of the NhaA Na+/H+ Exchanger of E. coli 166 6.5.2 Cotransport: A Sugar-Induced Electrogenic Partial Reaction in the Lactose Permease LacY of E. coli 168 6.5.3 The Glutamate Transporter EAAC1: A Robust Electrophysiological Assay with High Information Content 170 6.6 Conclusions 172 References 173 7 Stopped - Flow Fluorimetry Using Voltage - Sensitive Fluorescent Membrane Probes 179; Ronald J. Clarke and Mohammed A. A. Khalid 7.1 Introduction 179 7.2 Basics of the Stopped-Flow Technique 181 7.2.1 Flow Cell Design 181 7.2.2 Rapid Data Acquisition 181 7.2.3 Dead Time 183
• Edition: 1st
• Publisher Details: Hoboken, New Jersey : John Wiley & Sons
• Publication Date: 2015
• Extent: 1 online resource
• Subjects: 572.696; Carrier proteins; Ion pumps; Ion channels; Membrane proteins; Biological transport
• Languages: English
• ISBNs: 9781119085140
• Related ISBNs: 9781119085133
• Notes: Note: Description based on CIP data; resource not viewed.
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