Expression and Analysis of Recombinant Ion Channels is directed at both the new and the experienced researcher wishing to learn more about the considerations and methods for studying recombinant ion channels.

Features:

• Contains contributions from authors who routinely apply these approaches in key academic labs or major pharmaceutical companies
• Provides a comprehensive and easily accessible overview of the techniques involved
• Reviews underlying principles and and providing working guidelines as well as an understanding of the key theoretical and practical considerations associated with each topic

Contents

Expression of Ion Channels in Xenopus Oocytes

• Advantages and Disadvantages of Xenopus Oocytes.
• Procedures for Using Oocytes.
• Types of Analyses.
• Examples of Use

Molecular Biology Techniques for Structure – Function Studies of Ion Channels

• Methods for cDNA Subcloning.
• Generation of Chimeric Channel cDNAs.
• Site-directed Mutagenesis.
• Epitope-tagged Channels and Fusion Partners.
• Channel Subunit Concatamers

Unnatural Amino Acids as Probes of Ion Channel Structure – Function and Pharmacology

• Unnatural Amino Acid Mutagenesis Methodology.
• Unnatural Amino Acid Mutagenesis for Ion Channel Studies.
• Structure–Function Example Studies.
• Other Uses of Unnatural Amino Acids as Probes of Protein Structure and Function

Functional Expression of Ion Channels in Mammalian Systems

• cDNA Cloning and Manipulation.
• Choice of Host Cell Background.
• Post-translational Processing of Heterologous Expressed Ion Channels.
• Cytotoxicity.
• Transient Expression Systems.
• Stable Expression of Ion ChannelsM

Analysis of Electrophysiological Data

• Expression Systems and Related Recording Techniques.
• Macroscopic Recordings.
• Single Channel Analysis

Automated Planar Array Electrophysiology for Ion Channel Research

• Overview of Planar Array Recording.
• Experimental Methods and Design.
• Overall Success Rates and Throughput.
• Population Patch Clamp

Ion Flux and Ligand Binding Assays for Analysisof Ion Channels

• Ion Flux Assays.
• Ligand Binding Assays

Ion Channel Assays Based on Ion and Voltage-sensitive Fluorescent Probes

• Membrane Potential Probes.
• Ion-sensitive Fluorescent Probes.
• Fluorescence Assays for Ion Channels.
• Assays for Monitoring Channel Trafficking

Approaches for Ion Channel Structural Studies

• Expression of Membrane Proteins for Structural Studies.
• The Detergent Factor.
• Purification.
• Crystallization.
• Use of Antibody Fragments.
• Generation of First Diffraction Datasets.
• Selenomethionine Phasing of Membrane Proteins.
• MAD Phasing and Edge Scanning.
• Negative B- factor Application (Structure Factor Sharpening)

Molecular Modeling and Simulations of Ion Channels: Applications to Potassium Channels

• Computational Methods.
• Kir Channels.
• Kv Channels.

Index

click here to see books • videos • cd-roms of related interest