Proton transport by halorhodopsin

Biochemistry. 1996 May 28;35(21):6604-11. doi: 10.1021/bi9601159.


In halorhodopsin from Natronobacterium pharaonis, a light-driven chloride pump, the chloride binding site also binds azide. When azide is bound at this location the retinal Schiff base transiently deprotonates after photoexcitation with light > 530 nm, like in the light-driven proton pump bacteriorhodopsin. As in the photocycle of bacteriorhodopsin, pyranine detects the release of protons to the bulk. The subsequent reprotonation of the Schiff base is also dependent on azide, but with different kinetics that suggest a shuttling of protons from the surface as described earlier for halorhodopsin from Halobacterium salinarium. This azide-dependent, bacteriorhodopsin-like photocycle results in active electrogenic proton transport in the cytoplasmic to extracellular direction, detected in cell envelope vesicle suspensions both with a potential-sensitive electrode and by measuring light-dependent pH change. We conclude that in halorhodopsin an azide bound to the extracellular side of the Schiff base, and another azide shuttling between the Schiff base and the cytoplasmic surface, fulfill the functions of Asp-85 and Asp-96, respectively, in bacteriorhodopsin. Thus, although halorhodopsin is normally a chloride ion pump, it evidently contains all structural requirements, except an internal proton acceptor and a donor, of a proton pump. This observation complements our earlier finding that when a chloride binding site was created in bacteriorhodopsin through replacement of Asp-85 with a threonine, that protein became a chloride ion pump.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Azides / metabolism
  • Bacteriorhodopsins / metabolism*
  • Binding Sites
  • Binding, Competitive
  • Biological Transport, Active
  • Chlorides / metabolism
  • Halobacteriaceae / metabolism
  • Halobacterium / metabolism
  • Halorhodopsins
  • Hydrogen-Ion Concentration
  • Kinetics
  • Light
  • Protons
  • Schiff Bases


  • Azides
  • Chlorides
  • Halorhodopsins
  • Protons
  • Schiff Bases
  • Bacteriorhodopsins