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Protein-protein interactions in AQP regulation – biophysical characterization of AQP0-CaM and AQP2-LIP5 complex formation

Abstract

Protein-protein interactions play important roles in regulating human aquaporins (AQP) by gating as well as trafficking. While structural and functional studies have provided detailed knowledge of AQP transport mechanism, selectivity as well as gating by conformational changes of loops or termini, the mechanism behind how protein-protein interactions control AQP-mediated water transport through cellular membranes remain poorly characterized. Here we explore the interaction between two human AQPs and regulatory proteins: the interaction between AQP0 and calmodulin, which mediates AQP0 gating, as well as the interaction between AQP2 and LIP5, which is involved in trafficking. Using microscale thermophoresis (MST) and fluorescence anisotropy, two methods that have the advantage of low sample consumption and detergent compatibility, we show that the interactions can be studied using both full-length AQPs and AQP peptides corresponding to the regulatory protein binding sites. However, full-length AQPs gave better reproducibility between methods and for the first time revealed that AQP0 binds CaM in a cooperative manner, which was not seen in experiments using peptides. Our study highlights that, while peptides are great tools for locating binding sites and pinpointing interacting residues, full-length proteins may give additional insights, such as binding mechanism, allostery and cooperativity, which are important parameters for understanding protein-protein interactions in the cellular context.

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Publication details

The article was received on 09 Mar 2018, accepted on 23 Apr 2018 and first published on 25 Apr 2018


Article type: Paper
DOI: 10.1039/C8FD00065D
Citation: Faraday Discuss., 2018, Accepted Manuscript
  • Open access: Creative Commons BY-NC license
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    Protein-protein interactions in AQP regulation – biophysical characterization of AQP0-CaM and AQP2-LIP5 complex formation

    S. Kreida, J. V. Roche, C. Olsson, S. Linse and S. Törnroth-Horsefield, Faraday Discuss., 2018, Accepted Manuscript , DOI: 10.1039/C8FD00065D

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