The synthesis of recombinant membrane proteins in yeast for structural studies

Sarah Routledge, Lina Mikaliunaite, Anjana Patel, Michelle Clare, Stephanie P. Cartwright, Zharain Bawa, Martin D.B. Wilks, Floren Low, David Hardy, Alice J. Rothnie, Roslyn M. Bill*

*Corresponding author for this work

Research output: Contribution to journalArticle

Abstract

Historically, recombinant membrane protein production has been a major challenge meaning that many fewer membrane protein structures have been published than those of soluble proteins. However, there has been a recent, almost exponential increase in the number of membrane protein structures being deposited in the Protein Data Bank. This suggests that empirical methods are now available that can ensure the required protein supply for these difficult targets. This review focuses on methods that are available for protein production in yeast, which is an important source of recombinant eukaryotic membrane proteins. We provide an overview of approaches to optimize the expression plasmid, host cell and culture conditions, as well as the extraction and purification of functional protein for crystallization trials in preparation for structural studies.
Original languageEnglish
Pages (from-to)26-37
Number of pages12
JournalMethods
Volume95
Early online date30 Sep 2015
DOIs
Publication statusPublished - 15 Feb 2016

Fingerprint

Recombinant Proteins
Yeast
Membrane Proteins
Yeasts
Proteins
Crystallization
Purification
Plasmids
Cell Culture Techniques
Databases

Bibliographical note

Funding: BBSRC (grants BB/I019960/1, BB/K013319/1, BB/L502194/1 and BB/L015846/1; and the Innovative Medicines Joint Undertaking (grant 115583 to the ND4BB ENABLE Consortium)

Keywords

  • membrane proteins
  • yeast
  • recombinant antigen
  • expression plasmid vector
  • promoter
  • SMALP
  • cholesterol

Cite this

Routledge, Sarah ; Mikaliunaite, Lina ; Patel, Anjana ; Clare, Michelle ; Cartwright, Stephanie P. ; Bawa, Zharain ; Wilks, Martin D.B. ; Low, Floren ; Hardy, David ; Rothnie, Alice J. ; Bill, Roslyn M. / The synthesis of recombinant membrane proteins in yeast for structural studies. In: Methods. 2016 ; Vol. 95. pp. 26-37.
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Routledge, S, Mikaliunaite, L, Patel, A, Clare, M, Cartwright, SP, Bawa, Z, Wilks, MDB, Low, F, Hardy, D, Rothnie, AJ & Bill, RM 2016, 'The synthesis of recombinant membrane proteins in yeast for structural studies', Methods, vol. 95, pp. 26-37. https://doi.org/10.1016/j.ymeth.2015.09.027

The synthesis of recombinant membrane proteins in yeast for structural studies. / Routledge, Sarah; Mikaliunaite, Lina; Patel, Anjana; Clare, Michelle; Cartwright, Stephanie P.; Bawa, Zharain; Wilks, Martin D.B.; Low, Floren; Hardy, David; Rothnie, Alice J.; Bill, Roslyn M.

In: Methods, Vol. 95, 15.02.2016, p. 26-37.

Research output: Contribution to journalArticle

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