Microbial expression systems for membrane proteins

Marvin V. Dilworth, Mathilde S. Piel, Kim E. Bettaney, Pikyee Ma, Ji Luo, David Sharples, David R. Poyner, Stephane R. Gross, Karine Moncoq, Peter J. F. Henderson, Bruno Miroux, Roslyn M. Bill

Research output: Contribution to journalArticle

Abstract

Despite many high-profile successes, recombinant membrane protein production remains a technical challenge; it is still the case that many fewer membrane protein structures have been published than those of soluble proteins. However, progress is being made because empirical methods have been developed to produce the required quantity and quality of these challenging targets. This review focuses on the microbial expression systems that are a key source of recombinant prokaryotic and eukaryotic membrane proteins for structural studies. We provide an overview of the host strains, tags and promoters that, in our experience, are most likely to yield protein suitable for structural and functional characterization. We also catalogue the detergents used for solubilization and crystallization studies of these proteins. Here, we emphasize a combination of practical methods, not necessarily high-throughput, which can be implemented in any laboratory equipped for recombinant DNA technology and microbial cell culture.
LanguageEnglish
JournalMethods
Early online date12 Apr 2018
DOIs
Publication statusE-pub ahead of print - 12 Apr 2018

Fingerprint

Membrane Proteins
Genetic engineering
Proteins
Recombinant DNA
Crystallization
Cell culture
Recombinant Proteins
Detergents
Cell Culture Techniques
Throughput
Technology

Bibliographical note

© 2018 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/BY/4.0/).

Funding: Biotechnology and Biological Sciences Research Council (BBSRC; via grants BB/N007417/1, BB/P025927/1 and BB/P022685/1 to RMB) and the Innovative Medicines Joint Undertaking under Grant Agreement number 115583 to the ND4BB ENABLE Consortium to RMB. EC Membrane Protein Consortium (E-MeP, FP6 LSHG-CT-2004-504601), the European Drug Initiative for Channels and Transporters (EDICT, FP7 Health-F4-2007-201924 and BBBSRC grant BB/D524832/1 to RMB and PJFH. MVD’s PhD studentship was funded by BBSRC iCASE BB/H016244/1 with Glycoform Ltd. PJFH is grateful to the Leverhulme Trust for an Emeritus Research Fellowship (Grant number EM-2014-045). BBSRC (MPSI BBS/B/14418), the Wellcome Trust (JIF 062164/Z/00/Z) and the University of Leeds. ‘Initiative d'Excellence’ program from the French State (Grant ‘DYNAMO’, ANR-11-LABEX-0011-01). PhD fellowship funded by the French Ministry of Research and Education.

Keywords

  • Recombinant membrane proteins
  • Expression plasmid vector
  • Tag
  • Promoter
  • Detergent

Cite this

Dilworth, M. V., Piel, M. S., Bettaney, K. E., Ma, P., Luo, J., Sharples, D., ... Bill, R. M. (2018). Microbial expression systems for membrane proteins. Methods. https://doi.org/10.1016/j.ymeth.2018.04.009
Dilworth, Marvin V. ; Piel, Mathilde S. ; Bettaney, Kim E. ; Ma, Pikyee ; Luo, Ji ; Sharples, David ; Poyner, David R. ; Gross, Stephane R. ; Moncoq, Karine ; J. F. Henderson, Peter ; Miroux, Bruno ; Bill, Roslyn M. / Microbial expression systems for membrane proteins. In: Methods. 2018.
@article{e833462a252c4962a3c484c08a1c078a,
title = "Microbial expression systems for membrane proteins",
abstract = "Despite many high-profile successes, recombinant membrane protein production remains a technical challenge; it is still the case that many fewer membrane protein structures have been published than those of soluble proteins. However, progress is being made because empirical methods have been developed to produce the required quantity and quality of these challenging targets. This review focuses on the microbial expression systems that are a key source of recombinant prokaryotic and eukaryotic membrane proteins for structural studies. We provide an overview of the host strains, tags and promoters that, in our experience, are most likely to yield protein suitable for structural and functional characterization. We also catalogue the detergents used for solubilization and crystallization studies of these proteins. Here, we emphasize a combination of practical methods, not necessarily high-throughput, which can be implemented in any laboratory equipped for recombinant DNA technology and microbial cell culture.",
keywords = "Recombinant membrane proteins, Expression plasmid vector, Tag, Promoter, Detergent",
author = "Dilworth, {Marvin V.} and Piel, {Mathilde S.} and Bettaney, {Kim E.} and Pikyee Ma and Ji Luo and David Sharples and Poyner, {David R.} and Gross, {Stephane R.} and Karine Moncoq and {J. F. Henderson}, Peter and Bruno Miroux and Bill, {Roslyn M.}",
note = "{\circledC} 2018 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/BY/4.0/). Funding: Biotechnology and Biological Sciences Research Council (BBSRC; via grants BB/N007417/1, BB/P025927/1 and BB/P022685/1 to RMB) and the Innovative Medicines Joint Undertaking under Grant Agreement number 115583 to the ND4BB ENABLE Consortium to RMB. EC Membrane Protein Consortium (E-MeP, FP6 LSHG-CT-2004-504601), the European Drug Initiative for Channels and Transporters (EDICT, FP7 Health-F4-2007-201924 and BBBSRC grant BB/D524832/1 to RMB and PJFH. MVD’s PhD studentship was funded by BBSRC iCASE BB/H016244/1 with Glycoform Ltd. PJFH is grateful to the Leverhulme Trust for an Emeritus Research Fellowship (Grant number EM-2014-045). BBSRC (MPSI BBS/B/14418), the Wellcome Trust (JIF 062164/Z/00/Z) and the University of Leeds. ‘Initiative d'Excellence’ program from the French State (Grant ‘DYNAMO’, ANR-11-LABEX-0011-01). PhD fellowship funded by the French Ministry of Research and Education.",
year = "2018",
month = "4",
day = "12",
doi = "10.1016/j.ymeth.2018.04.009",
language = "English",
journal = "Methods",
issn = "1046-2023",
publisher = "Academic Press Inc.",

}

Dilworth, MV, Piel, MS, Bettaney, KE, Ma, P, Luo, J, Sharples, D, Poyner, DR, Gross, SR, Moncoq, K, J. F. Henderson, P, Miroux, B & Bill, RM 2018, 'Microbial expression systems for membrane proteins' Methods. https://doi.org/10.1016/j.ymeth.2018.04.009

Microbial expression systems for membrane proteins. / Dilworth, Marvin V.; Piel, Mathilde S.; Bettaney, Kim E.; Ma, Pikyee; Luo, Ji; Sharples, David; Poyner, David R.; Gross, Stephane R.; Moncoq, Karine; J. F. Henderson, Peter; Miroux, Bruno; Bill, Roslyn M.

In: Methods, 12.04.2018.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Microbial expression systems for membrane proteins

AU - Dilworth, Marvin V.

AU - Piel, Mathilde S.

AU - Bettaney, Kim E.

AU - Ma, Pikyee

AU - Luo, Ji

AU - Sharples, David

AU - Poyner, David R.

AU - Gross, Stephane R.

AU - Moncoq, Karine

AU - J. F. Henderson, Peter

AU - Miroux, Bruno

AU - Bill, Roslyn M.

N1 - © 2018 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/BY/4.0/). Funding: Biotechnology and Biological Sciences Research Council (BBSRC; via grants BB/N007417/1, BB/P025927/1 and BB/P022685/1 to RMB) and the Innovative Medicines Joint Undertaking under Grant Agreement number 115583 to the ND4BB ENABLE Consortium to RMB. EC Membrane Protein Consortium (E-MeP, FP6 LSHG-CT-2004-504601), the European Drug Initiative for Channels and Transporters (EDICT, FP7 Health-F4-2007-201924 and BBBSRC grant BB/D524832/1 to RMB and PJFH. MVD’s PhD studentship was funded by BBSRC iCASE BB/H016244/1 with Glycoform Ltd. PJFH is grateful to the Leverhulme Trust for an Emeritus Research Fellowship (Grant number EM-2014-045). BBSRC (MPSI BBS/B/14418), the Wellcome Trust (JIF 062164/Z/00/Z) and the University of Leeds. ‘Initiative d'Excellence’ program from the French State (Grant ‘DYNAMO’, ANR-11-LABEX-0011-01). PhD fellowship funded by the French Ministry of Research and Education.

PY - 2018/4/12

Y1 - 2018/4/12

N2 - Despite many high-profile successes, recombinant membrane protein production remains a technical challenge; it is still the case that many fewer membrane protein structures have been published than those of soluble proteins. However, progress is being made because empirical methods have been developed to produce the required quantity and quality of these challenging targets. This review focuses on the microbial expression systems that are a key source of recombinant prokaryotic and eukaryotic membrane proteins for structural studies. We provide an overview of the host strains, tags and promoters that, in our experience, are most likely to yield protein suitable for structural and functional characterization. We also catalogue the detergents used for solubilization and crystallization studies of these proteins. Here, we emphasize a combination of practical methods, not necessarily high-throughput, which can be implemented in any laboratory equipped for recombinant DNA technology and microbial cell culture.

AB - Despite many high-profile successes, recombinant membrane protein production remains a technical challenge; it is still the case that many fewer membrane protein structures have been published than those of soluble proteins. However, progress is being made because empirical methods have been developed to produce the required quantity and quality of these challenging targets. This review focuses on the microbial expression systems that are a key source of recombinant prokaryotic and eukaryotic membrane proteins for structural studies. We provide an overview of the host strains, tags and promoters that, in our experience, are most likely to yield protein suitable for structural and functional characterization. We also catalogue the detergents used for solubilization and crystallization studies of these proteins. Here, we emphasize a combination of practical methods, not necessarily high-throughput, which can be implemented in any laboratory equipped for recombinant DNA technology and microbial cell culture.

KW - Recombinant membrane proteins

KW - Expression plasmid vector

KW - Tag

KW - Promoter

KW - Detergent

UR - https://www.sciencedirect.com/science/article/pii/S1046202317303778?via%3Dihub

U2 - 10.1016/j.ymeth.2018.04.009

DO - 10.1016/j.ymeth.2018.04.009

M3 - Article

JO - Methods

T2 - Methods

JF - Methods

SN - 1046-2023

ER -

Dilworth MV, Piel MS, Bettaney KE, Ma P, Luo J, Sharples D et al. Microbial expression systems for membrane proteins. Methods. 2018 Apr 12. https://doi.org/10.1016/j.ymeth.2018.04.009