Membrane Protein Production in Insect Cells

Afroditi Vaitsopoulou; Peer Depping; Roslyn M Bill; Alan D Goddard; Alice J Rothnie

doi:10.1007/978-1-0716-2368-8_12

Membrane Protein Production in Insect Cells

Afroditi Vaitsopoulou, Peer Depping, Roslyn M Bill, Alan D Goddard, Alice J Rothnie

Research output: Chapter in Book/Published conference output › Chapter (peer-reviewed) › peer-review

Abstract

Membrane proteins are an essential part of the machinery of life. They connect the interior and exterior of cells, play an important role in cell signaling and are responsible for the influx and efflux of nutrients and metabolites. For their structural and functional analysis high yields of correctly folded and modified protein are needed. Insect cells, such as Sf9 cells, have been one of the major expression hosts for eukaryotic membrane proteins in structural investigations during the last decade, as they are easier to handle than mammalian cells and provide more natural posttranslational modifications than microbial systems. Here we describe general techniques for establishing and maintaining insect cell cultures, the generation and amplification of recombinant baculovirus stocks using the flashBAC™ or Bac-to-Bac™ systems, membrane protein production, as well as the production of membrane preparations for extraction and purification experiments.

Original language	English
Title of host publication	Methods in Molecular Biology
Editors	I. Mus-Veteau
Publisher	Springer
Pages	223-240
Number of pages	18
Volume	2507
ISBN (Electronic)	978-1-0716-2368-8
ISBN (Print)	978-1-0716-2367-1
DOIs	https://doi.org/10.1007/978-1-0716-2368-8_12
Publication status	Published - 1 Jul 2022

Publication series

Name	Methods in Molecular Biology
Volume	2507
ISSN (Print)	1064-3745
ISSN (Electronic)	1940-6029

Bibliographical note

Funding Information:
We are grateful for funding from the European Union’s Horizon 2020 research and innovation programme under Marie Sklodowska-Curie grant agreement No. 847419 (MemTrain) as well as the ERACoBioTech MeMBrane project and BBSRC (BB/R02152X/1) to A.D.G, A.J.R, and R.M.B.

Keywords

Animals
Baculoviridae/genetics
Genetic Vectors
Insecta/metabolism
Mammals/metabolism
Membrane Proteins/genetics
Protein Processing, Post-Translational
Recombinant Proteins/genetics
Sf9 Cells
Spodoptera/metabolism
Recombinant baculovirus
Sf9 insect cells
Membrane protein production

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10.1007/978-1-0716-2368-8_12

Cite this

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abstract = "Membrane proteins are an essential part of the machinery of life. They connect the interior and exterior of cells, play an important role in cell signaling and are responsible for the influx and efflux of nutrients and metabolites. For their structural and functional analysis high yields of correctly folded and modified protein are needed. Insect cells, such as Sf9 cells, have been one of the major expression hosts for eukaryotic membrane proteins in structural investigations during the last decade, as they are easier to handle than mammalian cells and provide more natural posttranslational modifications than microbial systems. Here we describe general techniques for establishing and maintaining insect cell cultures, the generation and amplification of recombinant baculovirus stocks using the flashBAC{\texttrademark} or Bac-to-Bac{\texttrademark} systems, membrane protein production, as well as the production of membrane preparations for extraction and purification experiments.",

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author = "Afroditi Vaitsopoulou and Peer Depping and Bill, {Roslyn M} and Goddard, {Alan D} and Rothnie, {Alice J}",

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AU - Goddard, Alan D

AU - Rothnie, Alice J

N1 - Funding Information: We are grateful for funding from the European Union’s Horizon 2020 research and innovation programme under Marie Sklodowska-Curie grant agreement No. 847419 (MemTrain) as well as the ERACoBioTech MeMBrane project and BBSRC (BB/R02152X/1) to A.D.G, A.J.R, and R.M.B.

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N2 - Membrane proteins are an essential part of the machinery of life. They connect the interior and exterior of cells, play an important role in cell signaling and are responsible for the influx and efflux of nutrients and metabolites. For their structural and functional analysis high yields of correctly folded and modified protein are needed. Insect cells, such as Sf9 cells, have been one of the major expression hosts for eukaryotic membrane proteins in structural investigations during the last decade, as they are easier to handle than mammalian cells and provide more natural posttranslational modifications than microbial systems. Here we describe general techniques for establishing and maintaining insect cell cultures, the generation and amplification of recombinant baculovirus stocks using the flashBAC™ or Bac-to-Bac™ systems, membrane protein production, as well as the production of membrane preparations for extraction and purification experiments.

AB - Membrane proteins are an essential part of the machinery of life. They connect the interior and exterior of cells, play an important role in cell signaling and are responsible for the influx and efflux of nutrients and metabolites. For their structural and functional analysis high yields of correctly folded and modified protein are needed. Insect cells, such as Sf9 cells, have been one of the major expression hosts for eukaryotic membrane proteins in structural investigations during the last decade, as they are easier to handle than mammalian cells and provide more natural posttranslational modifications than microbial systems. Here we describe general techniques for establishing and maintaining insect cell cultures, the generation and amplification of recombinant baculovirus stocks using the flashBAC™ or Bac-to-Bac™ systems, membrane protein production, as well as the production of membrane preparations for extraction and purification experiments.

KW - Animals

KW - Baculoviridae/genetics

KW - Genetic Vectors

KW - Insecta/metabolism

KW - Mammals/metabolism

KW - Membrane Proteins/genetics

KW - Protein Processing, Post-Translational

KW - Recombinant Proteins/genetics

KW - Sf9 Cells

KW - Spodoptera/metabolism

KW - Recombinant baculovirus

KW - Sf9 insect cells

KW - Membrane protein production

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C2 - 35773585

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BT - Methods in Molecular Biology

A2 - Mus-Veteau, I.

PB - Springer

ER -

Membrane Protein Production in Insect Cells

Abstract

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Bibliographical note

Keywords

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