Structural Lipids Enable the Formation of Functional Oligomers of the Eukaryotic Purine Symporter UapA. Issue 7 (19th July 2018)

• Record Type: Journal Article
• Authors: Pyle, Euan; Kalli, Antreas C.; Amillis, Sotiris; Hall, Zoe; Lau, Andy M.; Hanyaloglu, Aylin C.; Diallinas, George; Byrne, Bernadette; Politis, Argyris
• Is Part Of: Cell chemical biology. Volume 25:Issue 7(2018)
• Page Start: 840
• Page End: 848.e4
• Publication Date: 2018-07-19
• Subjects: protein-lipid interactions -- native mass spectrometry -- liquid chromatography-mass spectrometry -- lipidomics -- molecular dynamics simulations -- ion-mobility mass spectrometry -- UapA -- eukaryotic membrane protein -- in vivo mutational analyses -- membrane protein oligomerization; Biochemistry -- Periodicals; 572.05
• DOI: 10.1016/j.chembiol.2018.03.011
• Languages: English
• ISSNs: 2451-9456

Abstract

Summary: The role of membrane lipids in modulating eukaryotic transporter assembly and function remains unclear. We investigated the effect of membrane lipids in the structure and transport activity of the purine transporter UapA from Aspergillus nidulans . We found that UapA exists mainly as a dimer and that two lipid molecules bind per UapA dimer. We identified three phospholipid classes that co-purified with UapA: phosphatidylcholine, phosphatidylethanolamine (PE), and phosphatidylinositol (PI). UapA delipidation caused dissociation of the dimer into monomers. Subsequent addition of PI or PE rescued the UapA dimer and allowed recovery of bound lipids, suggesting a central role of these lipids in stabilizing the dimer. Molecular dynamics simulations predicted a lipid binding site near the UapA dimer interface. Mutational analyses established that lipid binding at this site is essential for formation of functional UapA dimers. We propose that structural lipids have a central role in the formation of functional, dimeric UapA. Graphical Abstract: Highlights: Mass spectrometry reveals specific lipid binding to the eukaryotic transporter UapA Interfacial lipids stabilize the functional UapA dimer MD simulations reveal the lipid binding sites Mutagenesis of a lipid binding site disrupts UapA dimerization and function in vivo Abstract : We describe the first in-depth analysis of membrane lipid interactions with a eukaryotic transporter using native mass spectrometry. We demonstrate that the binding of structural lipids is essential to maintain the stability of the functional UapA dimer in both the gas phase and in vivo .