Electrogenic Cation Binding in the Electroneutral Na+/H+ Antiporter of Pyrococcus abyssi.
Na+/H+antiporters in the CPA1 branch of the cation proton antiporter family drive the electroneutral exchange of H+against Na+ions and ensure pH homeostasis in eukaryotic and prokaryotic organisms. Although their transport cycle is overall electroneutral, specific partial reactions are electrogenic. Here, we present an electrophysiological study of the PaNhaP Na+/H+antiporter from Pyrococcus abyssi reconstituted into liposomes. Positive transient currents were recorded upon addition of Na+to PaNhaP proteoliposomes, indicating a reaction where positive charge is rapidly displaced into the proteoliposomes with a rate constant of k >200 s-1We attribute the recorded currents to an electrogenic reaction that includes Na+binding and possibly occlusion. Subsequently, positive charge is transported out of the cell associated with H+binding, so that the overall reaction is electroneutral. We show that the differences in pH profile and Na+affinity of PaNhaP and the related MjNhaP1 from Methanocaldococcus jannaschii can be attributed to an additional negatively charged glutamate residue in PaNhaP. The results are discussed in the context of the physiological function of PaNhaP and other microbial Na+/H+exchangers. We propose that both, electroneutral and electrogenic Na+/H+antiporters, represent a carefully tuned self-regulatory system, which drives the cytoplasmic pH back to neutral after any deviation.
From the Departments of Biological Chemistry and.|the Department of Biophysics, "Carol Davila" University of Medicine and Pharmacy, 050474 Bucharest, Romania.
Octavian
O
Călinescu
From the Departments of Biological Chemistry and.|Structural Biology, Max Planck Institute of Biophysics, 60438 Frankfurt am Main, Germany and.
Mark
M
Linder
Structural Biology, Max Planck Institute of Biophysics, 60438 Frankfurt am Main, Germany and.
David
D
Wöhlert
Structural Biology, Max Planck Institute of Biophysics, 60438 Frankfurt am Main, Germany and.
Özkan
Ö
Yildiz
Structural Biology, Max Planck Institute of Biophysics, 60438 Frankfurt am Main, Germany and.
Werner
W
Kühlbrandt
From the Departments of Biological Chemistry and fendler@biophys.mpg.de.
Klaus
http://orcid.org/0000-0002-5699-7060
K
Fendler
D019843:Archaeal Proteins / Q000378:metabolism*; D002412:Cations / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport; D043903:Pyrococcus abyssi / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*; D013379:Substrate Specificity
D019843:Archaeal Proteins; D002412:Cations; D017923:Sodium-Hydrogen Exchangers
PaNhaP; archaea; electrophysiology; membrane transport; pH regulation; sodium-proton exchange; solid supported membrane