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  Proton transfer dynamics on the surface of the late M state of bacteriorhodopsin

Nachliel, E., Gutman, M., Tittor, J., & Oesterhelt, D. (2002). Proton transfer dynamics on the surface of the late M state of bacteriorhodopsin. Biophysical Journal, 83(1), 416-426.

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Genre: Journal Article
Alternative Title : Biophys. J.


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Nachliel, E., Author
Gutman, M., Author
Tittor, J.1, 2, Author              
Oesterhelt, D.1, Author              
1Oesterhelt, Dieter / Membrane Biochemistry, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565164              
2Conti, Elena / Structural Cell Biology, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565144              


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 Abstract: The cytoplasmic surface of the BR (initial) state of bacteriorhodopsin is characterized by a cluster of three carboxylates that function as a proton-collecting antenna. Systematic replacement of most of the surface carboxylates indicated that the cluster is made of D104, E161, and E234 (Checover, S., Y. Marantz, E. Nachliel, M. Gutman, M. Pfeiffer, J. Tittor, D. Oesterhelt, and N. Dencher. 2001. Biochemistry. 40:4281-4292), yet the BR state is a resting configuration; thus, its proton-collecting antenna can only indicate the presence of its role in the photo-intermediates where the protein is re-protonated by protons coming from the cytoplasmic matrix. In the present study we used the D96N and the triple (D96G/F171C/F219L) mutant for monitoring the proton-collecting properties of the protein in its late M state. The protein was maintained in a steady M state by continuous illumination and subjected to reversible pulse protonation caused by repeated excitation of pyranine present in the reaction mixture. The re- protonation dynamics of the pyranine anion was subjected to kinetic analysis, and the rate constants of the reaction of free protons with the surface groups and the proton exchange reactions between them were calculated. The reconstruction of the experimental signal indicated that the late M state of bacteriorhodopsin exhibits an efficient mechanism of proton delivery to the unoccupied-most basic-residue on its cytoplasmic surface (D38), which exceeds that of the BR configuration of the protein. The kinetic analysis was carried out in conjunction with the published structure of the M state (Sass, H., G. Buldt, R. Gessenich, D. Hehn, D. Neff, R. Schlesinger, J. Berendzen, and P. Ormos. 2000. Nature. 406:649- 653), the model that resolves most of the cytoplasmic surface. The combination of the kinetic analysis and the structural information led to identification of two proton-conducting tracks on the protein's surface that are funneling protons to D38. One track is made of the carboxylate moieties of residues D36 and E237, while the other is made of D102 and E232. In the late M state the carboxylates of both tracks are closer to D38 than in the BR (initial) state, accounting for a more efficient proton equilibration between the bulk and the protein's proton entrance channel. The triple mutant resembles in the kinetic properties of its proton conducting surface more the BR-M state than the initial state confirming structural similarities with the BR-M state and differences to the BR initial state.


Language(s): eng - English
 Dates: 2002-07
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: eDoc: 31375
ISI: 000176445800036
 Degree: -



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Title: Biophysical Journal
  Alternative Title : Biophys. J.
Source Genre: Journal
Publ. Info: -
Pages: - Volume / Issue: 83 (1) Sequence Number: - Start / End Page: 416 - 426 Identifier: ISSN: 0006-3495