Bacteriophage membrane protein P9 as a fusion partner for the efficient expression of membrane proteins in Escherichia coli

•A novel membrane protein expression system is reported.•It was used to produce GPCRs, transporters, and ion channels of human origin.•Of the 14 membrane proteins, eight were highly expressed, three were moderately expressed.•Seven could be purified after extraction with the mild detergent. Despite...

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Bibliographic Details
Published in:Protein expression and purification 2015-12, Vol.116, p.12-18
Main Authors: Jung, Yuna, Jung, Hyeim, Lim, Dongbin
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Bacteriophage phi 6 - chemistry
Bacteriophage phi 6 - genetics
Base Sequence
Cloning, Molecular
Detergents - chemistry
Escherichia coli - genetics
Fusion partner
G-protein-coupled receptor
Humans
Ion Channels - chemistry
Ion Channels - genetics
Ion Channels - isolation & purification
Membrane integration
Membrane protein
Molecular Sequence Data
Phage φ6
Plasmids - chemistry
Plasmids - genetics
Receptors, G-Protein-Coupled - chemistry
Receptors, G-Protein-Coupled - genetics
Receptors, G-Protein-Coupled - isolation & purification
Recombinant Fusion Proteins - chemistry
Recombinant Fusion Proteins - genetics
Recombinant Fusion Proteins - isolation & purification
Therapeutic antibody
Up-Regulation
Viral Proteins - chemistry
Viral Proteins - genetics
Viral Proteins - isolation & purification
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Summary:•A novel membrane protein expression system is reported.•It was used to produce GPCRs, transporters, and ion channels of human origin.•Of the 14 membrane proteins, eight were highly expressed, three were moderately expressed.•Seven could be purified after extraction with the mild detergent. Despite their important roles and economic values, studies of membrane proteins have been hampered by the difficulties associated with obtaining sufficient amounts of protein. Here, we report a novel membrane protein expression system that uses the major envelope protein (P9) of phage φ6 as an N-terminal fusion partner. Phage membrane protein P9 facilitated the synthesis of target proteins and their integration into the Escherichia coli cell membrane. This system was used to produce various multi-pass transmembrane proteins, including G-protein-coupled receptors, transporters, and ion channels of human origin. Green fluorescent protein fusion was used to confirm the correct folding of the expressed proteins. Of the 14 membrane proteins tested, eight were highly expressed, three were moderately expressed, and three were barely expressed in E. coli. Seven of the eight highly expressed proteins could be purified after extraction with the mild detergent lauryldimethylamine-oxide. Although a few proteins have previously been developed as fusion partners to augment membrane protein production, we believe that the major envelope protein P9 described here is better suited to the efficient expression of eukaryotic transmembrane proteins in E. coli.
ISSN:1046-5928
1096-0279
DOI:10.1016/j.pep.2015.07.010