Aquaporins (AQPs) are water channels embedded in the cell membrane that are critical in maintaining water homeostasis. We describe a protocol for determining the water permeation capacity of AQPs reconstituted into proteoliposomes. Using a stopped-flow setup, AQP embedded in proteoliposomes are exposed to an osmogenic gradient that triggers water flux. The consequent effects on proteoliposome size can be tracked using the fluorescence of an internalized fluorophore. This enables controlled characterization of water flux by AQPs. For complete details on the use and execution of this protocol, please refer to Kitchen et al. (2020). [Abstract copyright: © 2022 The Authors.]
|Early online date||13 Apr 2022|
|Publication status||Published - 17 Jun 2022|
Bibliographical note© 2022 The Authors. This is an open access article under the CC BY license (https://creativecommons.org/licenses/by/4.0/).
The authors acknowledge 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); the Swedish Research Council (to S.T-H. through 2013-05945 and 2019-06106); the Crafoord Foundation (to S.T.-H. through 20140811 and 20180916); the Magnus Bergvall Foundation (to S.T-H. through 2015-01534); 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 by Leverhulme Trust UK (ECF-2021-602 ).
S.T.H. P.G. and J.W.M. conceived the study and devised the experimental strategy. J.W.M. and J.H.S. performed the experiments and analyzed the data. All authors contributed to manuscript writing and approved the final version. The authors declare no competing interests.
- Water - metabolism
- Cell Membrane
- Single-molecule Assays
- Proteolipids - metabolism
- Aquaporins - metabolism
- Protein Biochemistry