title: Molecular impact of the membrane potential on the regulatory mechanism of proton transfer in sensory rhodopsin II.
abstract: Metabolism establishes a potential difference across the cell membrane of every living cell which drives and regulates secondary ion and solute transfer across membrane proteins. Unraveling the effect of the membrane potential on the level of single molecular groups of the membrane protein was long hampered by the lack of appropriate analytical techniques. We have developed Surface Enhanced Infrared Difference Absorption Spectroscopy (SEIDAS), a highly sensitive vibrational technique for surface analysis, for the study of solid-supported monolayers of orientated membrane proteins. Here, we present spectroscopic data on vibrational changes of sensory rhodopsin II from Natronomonas pharaonis (NpSR II). The application of the electrode potential provides a voltage drop across the NpSR II monolayer through the Helmholtz double layer that mimics the cellular membrane potential. IR difference spectra indicated a shift of the photostationary equilibrium from an M and O mixture toward an M dominant equilibrium. The shift of positive to negative potential exhibited similar effects on the light-induced SEIDA spectra as the increase in pH. This effect is explained in terms of local pH change raised by the compensation of excess charge from the electrode. As we have shown earlier (Jiang, et al. Proc. Natl. Acad. Sci. U.S.A. 2008, 105 (34), 12113-12117), the application of an electric field opposite to the physiological proton transfer from the retinal Schiff base to its counterion Asp75 leads to the selective halt of the latter. However, when the solution pH is much higher than 5.8, that is, when the proton releasing group at the extracellular side is ionized, proton transfer of Asp75 becomes insensitive to the electric field exerted by the electrode. We infer that the deprotonation of the proton release group creates a local polar environment surrounding Asp75 as a consequence of hydrogen-bonding rearrangements that exceeds the energy of the external dipole. Our results reveal a molecular model for the physiological regulation of the photocycle of NpSR II by the potential drop across the membrane which came about by the interplay between the change in local pH at the membrane surface and the external electric field.
authors.affiliation: Department of Chemistry, Biophysical Chemistry (PC III), Bielefeld University, 33615 Bielefeld, Germany.
authors.forename: Xiue
authors.initials: X
authors.lastname: Jiang
authors.forename: Martin
authors.initials: M
authors.lastname: Engelhard
authors.forename: Kenichi
authors.initials: K
authors.lastname: Ataka
authors.forename: Joachim
authors.initials: J
authors.lastname: Heberle
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry* / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D004563:Electrochemistry; D004566:Electrodes; D006216:Halobacteriaceae / Q000737:chemistry / Q000378:metabolism; D006860:Hydrogen Bonding; D006863:Hydrogen-Ion Concentration; D008564:Membrane Potentials / Q000502:physiology*; D008958:Models, Molecular; D017494:Proton Pumps / Q000737:chemistry* / Q000378:metabolism; D013055:Spectrophotometry, Infrared; D013499:Surface Properties; D014732:Vibration
chemical_list: D019843:Archaeal Proteins; C434451:HtrII protein, Natronobacterium pharaonis; D017494:Proton Pumps