Heterologous Expression of Membrane Proteins

Methods and Protocols
Third Auflage 2022
ISBN: 978-1-0716-2367-1
Verlag: Springer US

Buch, Englisch, Band 2507, 467 Seiten, Format (B × H): 183 mm x 260 mm, Gewicht: 1107 g

Reihe: Methods in Molecular Biology

This detailed volume explores protocols for the production of membrane proteins in a panel of heterologous organisms for structural studies. Beginning with techniques using E. coli as a host for the overproduction and purification of membrane proteins, the book continues with chapters covering mammalian membrane protein production in yeast, insect cells, mammalian cells, as well as using virus like particles and acellular systems. Additionally, new detergents and alternatives to detergents allowing membrane protein purification for structural analyses are described. The book closes with a chapter exploring the use of microscale thermophoresis (MST) to evaluate the binding activity of heterologously expressed proteins directly in crude membrane extracts. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratoryprotocols, and tips on troubleshooting and avoiding known pitfalls. 
Authoritative and up-to-date, Heterologous Expression of Membrane Proteins: Methods and Protocols, Third Edition serves as an ideal guide for scientists aiming to produce and purify functional recombinant membrane proteins for structural studies.

Zielgruppe

Professional/practitioner

Autoren/Hrsg.

Mus-Veteau, Isabelle

Fachgebiete

• Naturwissenschaften Biowissenschaften Zellbiologie
• Naturwissenschaften Chemie Chemie Allgemein

Inhaltsverzeichnis

Solubilization and Purification of abIntegrin from Rat Liver for Reconstitution into Nanodiscs.- Membrane Proteins Production in Escherichia coli: Protocols and Rules.- Functional Overexpression of Membrane Proteins in E. coli: The Good, the Bad, and the Ugly.- Heterologous Expression of Membrane Proteins in E. coli.- Overexpression of the ABC Transporter BmrA within Intracellular Caveolæ in Escherichia coli.- Production of Membrane Proteins in Pseudomonas stutzeri.- Overproduction of Membrane-Associated, and Integrated, Proteins Using Saccharomyces cerevisiae.- Purification of Membrane Proteins Overexpressed in Saccharomyces cerevisiae.- Production and Purification of a GFP-Tagged ABC Transporter CaCdr1p.- Membrane Protein Production in the Yeast, P. pastoris.- Production and Preparation of Isotopically-Labelled Human Membrane Proteins in Pichia pastoris for Fast-MAS-NMR Analyses.- Membrane Protein Production in Insect Cells.- Thromboxane AG Protein-Coupled Receptor Production and Crystallization for Structure Studies.- Production of Human ABC Transporters and Oligosaccharyltransferase Complexes for Structural Studies.- The Heterologous Expression and Purification of GPCRs.- Expression and Purification of the Human Thyroid-Stimulating Hormone Receptor.- Optimization of Recombinant GPCR Proteins for Biophysical and Structural Studies Using Virus-Like Particles.- Peptide Tags and Domains for Expression and Detection of Mammalian Membrane Proteins at the Cell Surface.- Tailor the Purification and Analysis of Membrane Proteins with Modular Detergents.- Detergent Alternatives: Membrane Protein Purification Using Synthetic Nanodisc Polymers.- Detergent-Free Membrane Protein Purification Using SMA Polymer.- Cell-Free Expression of GPCRs into Nanomembranes for Functional and Structural Studies.- Cell-Free Expression of Proton-Coupled Folate Transporter in the Presence of Nanodiscs.- Microscale Thermophoresis to Evaluate the Functionality of Heterologously Overexpressed Membrane Proteins in Membrane Preparations.