High-yield overproduction and purification of human aquaporins from Pichia pastoris

Tamim Al-Jubair; Jonas Hyld Steffen; Julie Winkel Missel; Philip Kitchen; Mootaz M. Salman; Roslyn M. Bill; Pontus Gourdon; Susanna Törnroth-Horsefield

doi:10.1016/j.xpro.2022.101298

High-yield overproduction and purification of human aquaporins from Pichia pastoris

Tamim Al-Jubair^*, Jonas Hyld Steffen, Julie Winkel Missel, Philip Kitchen, Mootaz M. Salman, Roslyn M. Bill, Pontus Gourdon, Susanna Törnroth-Horsefield

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

Abstract

Aquaporins (AQPs) are membrane-bound water channels that play crucial roles in maintaining the water homeostasis of the human body. Here, we present a protocol for high-yield recombinant expression of human AQPs in the methylotropic yeast Pichia pastoris and subsequent AQP purification. The protocol typically yields 1–5 mg AQP per g of yeast cell at >95% purity and is compatible with any membrane protein cloned into Pichia pastoris, although expression levels may vary. For complete details on the use and execution of this protocol, please refer to Kitchen et al. (2020) and Frick et al. (2014).

Original language	English
Article number	101298
Journal	STAR protocols
Volume	3
Issue number	2
Early online date	12 Apr 2022
DOIs	https://doi.org/10.1016/j.xpro.2022.101298
Publication status	Published - 17 Jun 2022

Bibliographical note

© 2022 The Authors. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

Funding Information:
We acknowledge funding from the Swedish Research Council (to S.T.-H. through 2013-05945 and 2019-06106 ), the Crafoord Foundation (to S.T.-H. through 20140711 and 20180916 ), the Magnus Bergvall Foundation (to S.T.-H. through 2015-01534 ), the Knut and Alice Wallenberg Foundation (to P.G. through KAW 2015.0131 and 2020.0194 ), the Lundbeck Foundation (to P.G. through R133-A12689 and R313-2019-774 ), the Independent Research Fund Denmark (to P.G. through 4183-00559 ), NordForsk (to P.G. through 82000 ), and UK Biotechnology & Biosciences Research Council (to R.M.B., and P.K. through BB/P025927/1 ). P.K. is the recipient of an Aston University 50th Anniversary Prize Fellowship; M.M.S. is supported through funding from Biogen to Prof. Tomas Kirchhausen.

Keywords

Cell Membrane
Molecular Biology
Protein Biochemistry
Protein expression and purification

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10.1016/j.xpro.2022.101298Licence: CC BY 4.0

High-yield overproduction and purification of human aquaporins from Pichia pastoris
© 2022 The Authors. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
Final published version, 2.42 MBLicence: CC BY 4.0

Cite this

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abstract = "Aquaporins (AQPs) are membrane-bound water channels that play crucial roles in maintaining the water homeostasis of the human body. Here, we present a protocol for high-yield recombinant expression of human AQPs in the methylotropic yeast Pichia pastoris and subsequent AQP purification. The protocol typically yields 1–5 mg AQP per g of yeast cell at >95% purity and is compatible with any membrane protein cloned into Pichia pastoris, although expression levels may vary. For complete details on the use and execution of this protocol, please refer to Kitchen et al. (2020) and Frick et al. (2014).",

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High-yield overproduction and purification of human aquaporins from Pichia pastoris

Abstract

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Keywords

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