On the track of the lipid transport pathway of the phospholipid flippase ATP8A2 - Mutation analysis of residues of the transmembrane segments M1, M2, M3 and M4.

Autor: Mogensen LS; Department of Biomedicine, Aarhus University, Aarhus, Denmark., Mikkelsen SA; Department of Biomedicine, Aarhus University, Aarhus, Denmark., Tadini-Buoninsegni F; Department of Chemistry 'Ugo Schiff', University of Florence, Sesto Fiorentino, Italy., Holm R; Department of Biomedicine, Aarhus University, Aarhus, Denmark., Matsell E; Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, Canada., Vilsen B; Department of Biomedicine, Aarhus University, Aarhus, Denmark., Molday RS; Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, Canada; Department of Ophthalmology and Visual Sciences, Centre for Macular Research, University of British Columbia, Vancouver, British Columbia, Canada., Andersen JP; Department of Biomedicine, Aarhus University, Aarhus, Denmark. Electronic address: jpa@biomed.au.dk.
Jazyk: angličtina
Zdroj: Biochimica et biophysica acta. Molecular cell research [Biochim Biophys Acta Mol Cell Res] 2024 Jan; Vol. 1871 (1), pp. 119570. Date of Electronic Publication: 2023 Sep 09.
DOI: 10.1016/j.bbamcr.2023.119570
Abstrakt: P4-ATPases, also known as flippases, translocate specific lipids from the exoplasmic leaflet to the cytoplasmic leaflet of biological membranes, thereby generating an asymmetric lipid distribution essential for numerous cellular functions. A debated issue is which pathway within the protein the lipid substrate follows during the translocation. Here we present a comprehensive mutational screening of all amino acid residues in the transmembrane segments M1, M2, M3, and M4 of the flippase ATP8A2, thus allowing the functionally important residues in these transmembrane segments to be highlighted on a background of less important residues. Kinetic analysis of ATPase activity of 130 new ATP8A2 mutants, providing V max values as well as apparent affinities of the mutants for the lipid substrate, support a translocation pathway between M2 and M4 ("M2-M4 path"), extending from the entry site, where the lipid substrate binds from the exoplasmic leaflet, to a putative exit site at the cytoplasmic surface, formed by the divergence of M2 and M4. The effects of mutations in the M2-M4 path on the function of the entry site, including loss of lipid specificity in some mutants, suggest that the M2-M4 path and the entry site are conformationally coupled. Many of the residues of the M2-M4 path possess side chains with a potential for interacting with each other in a zipper-like mode, as well as with the head group of the lipid substrate, by ionic/hydrogen bonds. Thus, the translocation of the lipid substrate toward the cytoplasmic bilayer leaflet is comparable to unzipping a zipper of salt bridges/hydrogen bonds.
Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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