Polyamine Transport Assay Using Reconstituted Yeast Membranes

Sarah Van Veen; Shaun Martin; Marleen Schuermans; Peter Vangheluwe

doi:10.21769/BioProtoc.3888

Polyamine Transport Assay Using Reconstituted Yeast Membranes

Bio Protoc. 2021 Jan 20;11(2):e3888. doi: 10.21769/BioProtoc.3888.

Authors

Sarah Van Veen¹, Shaun Martin¹, Marleen Schuermans¹, Peter Vangheluwe¹

Affiliation

¹ Department of Cellular and Molecular Medicine, KU Leuven, Leuven, Belgium.

Abstract

ATP13A2/PARK9 is a late endo-/lysosomal P5B transport ATPase that is associated with several neurodegenerative disorders. We recently characterized ATP13A2 as a lysosomal polyamine exporter, which sheds light on the molecular identity of the unknown mammalian polyamine transport system. Here, we describe step by step a protocol to measure radiolabeled polyamine transport in reconstituted vesicles from yeast cells overexpressing human ATP13A2. This protocol was developed as part of our recent publication (van Veen et al., 2020 ) and will be useful for characterizing the transport function of other putative polyamine transporters, such as isoforms of the P5B transport ATPases.

Keywords: ATP13A2; P5 ATPase; Polyamine; Reconstitution; Spermine; Transport assay; Yeast membranes.