Abstract
Vacuolar ATPases are multisubunit protein complexes that are indispensable for acidification and pH homeostasis in a variety of physiological processes in all eukaryotic cells. An arginine residue (Arg735) in transmembrane helix 7 (TM7) of subunit a of the yeast ATPase is known to be essential for proton translocation. However, the specific mechanism of its involvement in proton transport remains to be determined. Arginine residues are usually assumed to "snorkel" toward the protein surface when exposed to a hydrophobic environment. Here, using solution NMR spectroscopy, molecular dynamics simulations, and in vivo yeast assays, we obtained evidence for the formation of a transient, membrane-embedded cation-π interaction in TM7 between Arg735 and two highly conserved nearby aromatic residues, Tyr733 and Trp737 We propose a mechanism by which the transient, membrane-embedded cation-π complex provides the necessary energy to keep the charged side chain of Arg735 within the hydrophobic membrane. Such cation-π interactions may define a general mechanism to retain charged amino acids in a hydrophobic membrane environment.
Highlights
Vacuolar ATPases are multisubunit protein complexes that are indispensable for acidification and pH homeostasis in a variety of physiological processes in all eukaryotic cells
Molecular dynamics (MD) simulations have revealed that a charged arginine in a transmembrane model would experience a large free energy barrier of ϳ 17 kcal/mol to move across a lipid bilayer [24]
In a previous study of transmembrane helix 7 (TM7) of V-ATPase in membrane mimetics, we found evidence of Arg735 being located in the hydrophobic environment [34] rather than near the polar surface, as typically found for membrane-bound peptides
Summary
A cation– interaction in a transmembrane helix of vacuolar ATPase retains the proton-transporting arginine in a hydrophobic environment. Hofbauer¶1, Florian Sarkleti¶, Martina Setzʈ, Nina Gubensak‡, Sabine Lichtenegger§, Salvatore Fabio Falsone**, Heimo Wolinski¶, Simone Kosol‡‡, Chris Oostenbrinkʈ, Sepp D. Kohlwein¶, and X Klaus Zangger‡2 From the Institutes of ‡Chemistry and **Pharmaceutical Sciences, University of Graz, 8010 Graz, Austria, the §Institute of Hygiene, Microbiology, and Environmental Medicine, Medical University of Graz, 8010 Graz, Austria, the ¶Institute of Molecular Biosciences, BioTechMed-Graz, University of Graz, 8010 Graz, Austria, the ʈInstitute of Molecular Modeling and Simulation, University of Natural Resources and Life Sciences, 1190 Vienna, Austria, the ‡‡Department of Chemistry, University of Warwick, Gibbet Hill, Coventry CV4 7AL, United Kingdom
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