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. 2002 Jul;83(1):416–426. doi: 10.1016/S0006-3495(02)75179-5

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

Esther Nachliel 1, Menachem Gutman 1, Jörg Tittor 1, Dieter Oesterhelt 1
PMCID: PMC1302157  PMID: 12080130

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. Büldt, 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.

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Selected References

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  1. Brandsburg-Zabary S., Fried O., Marantz Y., Nachliel E., Gutman M. Biophysical aspects of intra-protein proton transfer. Biochim Biophys Acta. 2000 May 12;1458(1):120–134. doi: 10.1016/s0005-2728(00)00063-3. [DOI] [PubMed] [Google Scholar]
  2. Brown L. S., Dioumaev A. K., Needleman R., Lanyi J. K. Local-access model for proton transfer in bacteriorhodopsin. Biochemistry. 1998 Mar 17;37(11):3982–3993. doi: 10.1021/bi9728396. [DOI] [PubMed] [Google Scholar]
  3. Brown L. S., Sasaki J., Kandori H., Maeda A., Needleman R., Lanyi J. K. Glutamic acid 204 is the terminal proton release group at the extracellular surface of bacteriorhodopsin. J Biol Chem. 1995 Nov 10;270(45):27122–27126. doi: 10.1074/jbc.270.45.27122. [DOI] [PubMed] [Google Scholar]
  4. Cao Y., Váró G., Chang M., Ni B. F., Needleman R., Lanyi J. K. Water is required for proton transfer from aspartate-96 to the bacteriorhodopsin Schiff base. Biochemistry. 1991 Nov 12;30(45):10972–10979. doi: 10.1021/bi00109a023. [DOI] [PubMed] [Google Scholar]
  5. Checover S., Marantz Y., Nachliel E., Gutman M., Pfeiffer M., Tittor J., Oesterhelt D., Dencher N. A. Dynamics of the proton transfer reaction on the cytoplasmic surface of bacteriorhodopsin. Biochemistry. 2001 Apr 10;40(14):4281–4292. doi: 10.1021/bi002574m. [DOI] [PubMed] [Google Scholar]
  6. Checover S., Nachliel E., Dencher N. A., Gutman M. Mechanism of proton entry into the cytoplasmic section of the proton-conducting channel of bacteriorhodopsin. Biochemistry. 1997 Nov 11;36(45):13919–13928. doi: 10.1021/bi9717542. [DOI] [PubMed] [Google Scholar]
  7. Chon Y. S., Sasaki J., Kandori H., Brown L. S., Lanyi J. K., Needleman R., Maeda A. Hydration of the counterion of the Schiff base in the chloride-transporting mutant of bacteriorhodopsin: FTIR and FT-raman studies of the effects of anion binding when Asp85 is replaced with a neutral residue. Biochemistry. 1996 Nov 12;35(45):14244–14250. doi: 10.1021/bi9606197. [DOI] [PubMed] [Google Scholar]
  8. Dencher N. A., Sass H. J., Büldt G. Water and bacteriorhodopsin: structure, dynamics, and function. Biochim Biophys Acta. 2000 Aug 30;1460(1):192–203. doi: 10.1016/s0005-2728(00)00139-0. [DOI] [PubMed] [Google Scholar]
  9. Gutman M. Application of the laser-induced proton pulse for measuring the protonation rate constants of specific sites on proteins and membranes. Methods Enzymol. 1986;127:522–538. doi: 10.1016/0076-6879(86)27042-1. [DOI] [PubMed] [Google Scholar]
  10. Gutman M., Nachliel E., Gershon E. Effect of buffer on kinetics of proton equilibration with a protonable group. Biochemistry. 1985 Jun 4;24(12):2937–2941. doi: 10.1021/bi00333a019. [DOI] [PubMed] [Google Scholar]
  11. Gutman M. The pH jump: probing of macromolecules and solutions by a laser-induced, ultrashort proton pulse--theory and applications in biochemistry. Methods Biochem Anal. 1984;30:1–103. doi: 10.1002/9780470110515.ch1. [DOI] [PubMed] [Google Scholar]
  12. Hatanaka M., Sasaki J., Kandori H., Ebrey T. G., Needleman R., Lanyi J. K., Maeda A. Effects of arginine-82 on the interactions of internal water molecules in bacteriorhodopsin. Biochemistry. 1996 May 21;35(20):6308–6312. doi: 10.1021/bi952973v. [DOI] [PubMed] [Google Scholar]
  13. Haupts U., Bamberg E., Oesterhelt D. Different modes of proton translocation by sensory rhodopsin I. EMBO J. 1996 Apr 15;15(8):1834–1841. [PMC free article] [PubMed] [Google Scholar]
  14. Haupts U., Tittor J., Bamberg E., Oesterhelt D. General concept for ion translocation by halobacterial retinal proteins: the isomerization/switch/transfer (IST) model. Biochemistry. 1997 Jan 7;36(1):2–7. doi: 10.1021/bi962014g. [DOI] [PubMed] [Google Scholar]
  15. Heberle J., Dencher N. A. Surface-bound optical probes monitor protein translocation and surface potential changes during the bacteriorhodopsin photocycle. Proc Natl Acad Sci U S A. 1992 Jul 1;89(13):5996–6000. doi: 10.1073/pnas.89.13.5996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Heberle J., Oesterhelt D., Dencher N. A. Decoupling of photo- and proton cycle in the Asp85-->Glu mutant of bacteriorhodopsin. EMBO J. 1993 Oct;12(10):3721–3727. doi: 10.1002/j.1460-2075.1993.tb06049.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Heberle J, Fitter J, Sass HJ, Buldt G. Bacteriorhodopsin: the functional details of a molecular machine are being resolved. Biophys Chem. 2000 Jul 15;85(2-3):229–248. doi: 10.1016/s0301-4622(99)00154-4. [DOI] [PubMed] [Google Scholar]
  18. Henderson R., Baldwin J. M., Ceska T. A., Zemlin F., Beckmann E., Downing K. H. Model for the structure of bacteriorhodopsin based on high-resolution electron cryo-microscopy. J Mol Biol. 1990 Jun 20;213(4):899–929. doi: 10.1016/S0022-2836(05)80271-2. [DOI] [PubMed] [Google Scholar]
  19. Kandori H., Kinoshita N., Yamazaki Y., Maeda A., Shichida Y., Needleman R., Lanyi J. K., Bizounok M., Herzfeld J., Raap J. Structural change of threonine 89 upon photoisomerization in bacteriorhodopsin as revealed by polarized FTIR spectroscopy. Biochemistry. 1999 Jul 27;38(30):9676–9683. doi: 10.1021/bi990713y. [DOI] [PubMed] [Google Scholar]
  20. Koch M. H., Dencher N. A., Oesterhelt D., Plöhn H. J., Rapp G., Büldt G. Time-resolved X-ray diffraction study of structural changes associated with the photocycle of bacteriorhodopsin. EMBO J. 1991 Mar;10(3):521–526. doi: 10.1002/j.1460-2075.1991.tb07978.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lanyi J. K., Pohorille A. Proton pumps: mechanism of action and applications. Trends Biotechnol. 2001 Apr;19(4):140–144. doi: 10.1016/s0167-7799(01)01576-1. [DOI] [PubMed] [Google Scholar]
  22. Lanyi J. K. Progress toward an explicit mechanistic model for the light-driven pump, bacteriorhodopsin. FEBS Lett. 1999 Dec 31;464(3):103–107. doi: 10.1016/s0014-5793(99)01685-3. [DOI] [PubMed] [Google Scholar]
  23. Lanyi J. K. Understanding structure and function in the light-driven proton pump bacteriorhodopsin. J Struct Biol. 1998 Dec 15;124(2-3):164–178. doi: 10.1006/jsbi.1998.4044. [DOI] [PubMed] [Google Scholar]
  24. Luecke H., Schobert B., Richter H. T., Cartailler J. P., Lanyi J. K. Structural changes in bacteriorhodopsin during ion transport at 2 angstrom resolution. Science. 1999 Oct 8;286(5438):255–261. doi: 10.1126/science.286.5438.255. [DOI] [PubMed] [Google Scholar]
  25. Luecke H., Schobert B., Richter H. T., Cartailler J. P., Lanyi J. K. Structure of bacteriorhodopsin at 1.55 A resolution. J Mol Biol. 1999 Aug 27;291(4):899–911. doi: 10.1006/jmbi.1999.3027. [DOI] [PubMed] [Google Scholar]
  26. Mollaaghababa R., Steinhoff H. J., Hubbell W. L., Khorana H. G. Time-resolved site-directed spin-labeling studies of bacteriorhodopsin: loop-specific conformational changes in M. Biochemistry. 2000 Feb 8;39(5):1120–1127. doi: 10.1021/bi991963h. [DOI] [PubMed] [Google Scholar]
  27. Muneyuki E., Shibazaki C., Ohtani H., Okuno D., Asaumi M., Mogi T. Time-resolved measurements of photovoltage generation by bacteriorhodopsin and halorhodopsin adsorbed on a thin polymer film. J Biochem. 1999 Feb;125(2):270–276. doi: 10.1093/oxfordjournals.jbchem.a022283. [DOI] [PubMed] [Google Scholar]
  28. Müller D. J., Büldt G., Engel A. Force-induced conformational change of bacteriorhodopsin. J Mol Biol. 1995 Jun 2;249(2):239–243. doi: 10.1006/jmbi.1995.0292. [DOI] [PubMed] [Google Scholar]
  29. Nachliel E., Finkelstein Y., Gutman M. The mechanism of monensin-mediated cation exchange based on real time measurements. Biochim Biophys Acta. 1996 Dec 4;1285(2):131–145. doi: 10.1016/s0005-2736(96)00149-6. [DOI] [PubMed] [Google Scholar]
  30. Nachliel E., Gutman M., Kiryati S., Dencher N. A. Protonation dynamics of the extracellular and cytoplasmic surface of bacteriorhodopsin in the purple membrane. Proc Natl Acad Sci U S A. 1996 Oct 1;93(20):10747–10752. doi: 10.1073/pnas.93.20.10747. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nachliel E., Gutman M. Quantitative evaluation of the dynamics of proton transfer from photoactivated bacteriorhodopsin to the bulk. FEBS Lett. 1996 Sep 16;393(2-3):221–225. doi: 10.1016/0014-5793(96)00870-8. [DOI] [PubMed] [Google Scholar]
  32. Nagel G., Kelety B., Möckel B., Büldt G., Bamberg E. Voltage dependence of proton pumping by bacteriorhodopsin is regulated by the voltage-sensitive ratio of M1 to M2. Biophys J. 1998 Jan;74(1):403–412. doi: 10.1016/S0006-3495(98)77797-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Oesterhelt D. The structure and mechanism of the family of retinal proteins from halophilic archaea. Curr Opin Struct Biol. 1998 Aug;8(4):489–500. doi: 10.1016/s0959-440x(98)80128-0. [DOI] [PubMed] [Google Scholar]
  34. Oesterhelt D., Tittor J., Bamberg E. A unifying concept for ion translocation by retinal proteins. J Bioenerg Biomembr. 1992 Apr;24(2):181–191. doi: 10.1007/BF00762676. [DOI] [PubMed] [Google Scholar]
  35. Oesterhelt F., Oesterhelt D., Pfeiffer M., Engel A., Gaub H. E., Müller D. J. Unfolding pathways of individual bacteriorhodopsins. Science. 2000 Apr 7;288(5463):143–146. doi: 10.1126/science.288.5463.143. [DOI] [PubMed] [Google Scholar]
  36. Oka T., Kamikubo H., Tokunaga F., Lanyi J. K., Needleman R., Kataoka M. Conformational change of helix G in the bacteriorhodopsin photocycle: investigation with heavy atom labeling and x-ray diffraction. Biophys J. 1999 Feb;76(2):1018–1023. doi: 10.1016/S0006-3495(99)77266-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Otto H., Marti T., Holz M., Mogi T., Lindau M., Khorana H. G., Heyn M. P. Aspartic acid-96 is the internal proton donor in the reprotonation of the Schiff base of bacteriorhodopsin. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9228–9232. doi: 10.1073/pnas.86.23.9228. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pebay-Peyroula E., Rummel G., Rosenbusch J. P., Landau E. M. X-ray structure of bacteriorhodopsin at 2.5 angstroms from microcrystals grown in lipidic cubic phases. Science. 1997 Sep 12;277(5332):1676–1681. doi: 10.1126/science.277.5332.1676. [DOI] [PubMed] [Google Scholar]
  39. Pfeiffer M., Rink T., Gerwert K., Oesterhelt D., Steinhoff H. J. Site-directed spin-labeling reveals the orientation of the amino acid side-chains in the E-F loop of bacteriorhodopsin. J Mol Biol. 1999 Mar 19;287(1):163–171. doi: 10.1006/jmbi.1998.2593. [DOI] [PubMed] [Google Scholar]
  40. Radionov A. N., Kaulen A. D. Inhibition of the M1-->M2 (M(closed) --> M(open)) transition in the D96N mutant photocycle and its relation to the corresponding transition in wild-type bacteriorhodopsin. FEBS Lett. 1997 Jun 9;409(2):137–140. doi: 10.1016/s0014-5793(97)00474-2. [DOI] [PubMed] [Google Scholar]
  41. Radionov A. N., Kaulen A. D. Two forms of N intermediate (N(open) and N(closed)) in the bacteriorhodopsin photocycle. FEBS Lett. 1999 May 21;451(2):147–151. doi: 10.1016/s0014-5793(99)00577-3. [DOI] [PubMed] [Google Scholar]
  42. Radionov A. N., Klyachko V. A., Kaulen A. D. Formation of the M(N) (M(open)) intermediate in the wild-type bacteriorhodopsin photocycle is accompanied by an absorption spectrum shift to shorter wavelength, like that in the mutant D96N bacteriorhodopsin photocycle. Biochemistry (Mosc) 1999 Oct;64(10):1210–1214. [PubMed] [Google Scholar]
  43. Richter H. T., Needleman R., Lanyi J. K. Perturbed interaction between residues 85 and 204 in Tyr-185-->Phe and Asp-85-->Glu bacteriorhodopsins. Biophys J. 1996 Dec;71(6):3392–3398. doi: 10.1016/S0006-3495(96)79532-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Riesle J., Oesterhelt D., Dencher N. A., Heberle J. D38 is an essential part of the proton translocation pathway in bacteriorhodopsin. Biochemistry. 1996 May 28;35(21):6635–6643. doi: 10.1021/bi9600456. [DOI] [PubMed] [Google Scholar]
  45. Royant A., Edman K., Ursby T., Pebay-Peyroula E., Landau E. M., Neutze R. Helix deformation is coupled to vectorial proton transport in the photocycle of bacteriorhodopsin. Nature. 2000 Aug 10;406(6796):645–648. doi: 10.1038/35020599. [DOI] [PubMed] [Google Scholar]
  46. Rödig C., Siebert F. Distortion of the L-->M transition in the photocycle of the bacteriorhodopsin mutant D96N: a time-resolved step-scan FTIR investigation. FEBS Lett. 1999 Feb 19;445(1):14–18. doi: 10.1016/s0014-5793(99)00088-5. [DOI] [PubMed] [Google Scholar]
  47. Rüdiger M., Tittor J., Gerwert K., Oesterhelt D. Reconstitution of bacteriorhodopsin from the apoprotein and retinal studied by Fourier-transform infrared spectroscopy. Biochemistry. 1997 Apr 22;36(16):4867–4874. doi: 10.1021/bi962426p. [DOI] [PubMed] [Google Scholar]
  48. Sasaki J., Shichida Y., Lanyi J. K., Maeda A. Protein changes associated with reprotonation of the Schiff base in the photocycle of Asp96-->Asn bacteriorhodopsin. The MN intermediate with unprotonated Schiff base but N-like protein structure. J Biol Chem. 1992 Oct 15;267(29):20782–20786. [PubMed] [Google Scholar]
  49. Sass H. J., Büldt G., Gessenich R., Hehn D., Neff D., Schlesinger R., Berendzen J., Ormos P. Structural alterations for proton translocation in the M state of wild-type bacteriorhodopsin. Nature. 2000 Aug 10;406(6796):649–653. doi: 10.1038/35020607. [DOI] [PubMed] [Google Scholar]
  50. Sass H. J., Gessenich R., Koch M. H., Oesterhelt D., Dencher N. A., Büldt G., Rapp G. Evidence for charge-controlled conformational changes in the photocycle of bacteriorhodopsin. Biophys J. 1998 Jul;75(1):399–405. doi: 10.1016/S0006-3495(98)77524-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Sass H. J., Schachowa I. W., Rapp G., Koch M. H., Oesterhelt D., Dencher N. A., Büldt G. The tertiary structural changes in bacteriorhodopsin occur between M states: X-ray diffraction and Fourier transform infrared spectroscopy. EMBO J. 1997 Apr 1;16(7):1484–1491. doi: 10.1093/emboj/16.7.1484. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Steinhoff H. J., Pfeiffer M., Rink T., Burlon O., Kurz M., Riesle J., Heuberger E., Gerwert K., Oesterhelt D. Azide reduces the hydrophobic barrier of the bacteriorhodopsin proton channel. Biophys J. 1999 May;76(5):2702–2710. doi: 10.1016/S0006-3495(99)77422-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Steinhoff H., Savitsky A., Wegener C., Pfeiffer M., Plato M., Möbius K. High-field EPR studies of the structure and conformational changes of site-directed spin labeled bacteriorhodopsin. Biochim Biophys Acta. 2000 Apr 21;1457(3):253–262. doi: 10.1016/s0005-2728(00)00106-7. [DOI] [PubMed] [Google Scholar]
  54. Subramaniam S., Gerstein M., Oesterhelt D., Henderson R. Electron diffraction analysis of structural changes in the photocycle of bacteriorhodopsin. EMBO J. 1993 Jan;12(1):1–8. doi: 10.1002/j.1460-2075.1993.tb05625.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Subramaniam S., Lindahl M., Bullough P., Faruqi A. R., Tittor J., Oesterhelt D., Brown L., Lanyi J., Henderson R. Protein conformational changes in the bacteriorhodopsin photocycle. J Mol Biol. 1999 Mar 19;287(1):145–161. doi: 10.1006/jmbi.1999.2589. [DOI] [PubMed] [Google Scholar]
  56. Tanio M., Tuzi S., Yamaguchi S., Kawaminami R., Naito A., Needleman R., Lanyi J. K., Saitô H. Conformational changes of bacteriorhodopsin along the proton-conduction chain as studied with (13)C NMR of [3-(13)C]Ala-labeled protein: arg(82) may function as an information mediator. Biophys J. 1999 Sep;77(3):1577–1584. doi: 10.1016/S0006-3495(99)77005-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Thorgeirsson T. E., Xiao W., Brown L. S., Needleman R., Lanyi J. K., Shin Y. K. Transient channel-opening in bacteriorhodopsin: an EPR study. J Mol Biol. 1997 Nov 14;273(5):951–957. doi: 10.1006/jmbi.1997.1362. [DOI] [PubMed] [Google Scholar]
  58. Tittor J., Paula S., Subramaniam S., Heberle J., Henderson R., Oesterhelt D. Proton translocation by bacteriorhodopsin in the absence of substantial conformational changes. J Mol Biol. 2002 May 31;319(2):555–565. doi: 10.1016/S0022-2836(02)00307-8. [DOI] [PubMed] [Google Scholar]
  59. Tittor J., Schweiger U., Oesterhelt D., Bamberg E. Inversion of proton translocation in bacteriorhodopsin mutants D85N, D85T, and D85,96N. Biophys J. 1994 Oct;67(4):1682–1690. doi: 10.1016/S0006-3495(94)80642-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Váró G., Lanyi J. K. Effects of hydrostatic pressure on the kinetics reveal a volume increase during the bacteriorhodopsin photocycle. Biochemistry. 1995 Sep 26;34(38):12161–12169. doi: 10.1021/bi00038a009. [DOI] [PubMed] [Google Scholar]
  61. Váró G., Lanyi J. K. Kinetic and spectroscopic evidence for an irreversible step between deprotonation and reprotonation of the Schiff base in the bacteriorhodopsin photocycle. Biochemistry. 1991 May 21;30(20):5008–5015. doi: 10.1021/bi00234a024. [DOI] [PubMed] [Google Scholar]
  62. Váró G., Lanyi J. K. Protonation and deprotonation of the M, N, and O intermediates during the bacteriorhodopsin photocycle. Biochemistry. 1990 Jul 24;29(29):6858–6865. doi: 10.1021/bi00481a015. [DOI] [PubMed] [Google Scholar]
  63. Váró G., Lanyi J. K. Thermodynamics and energy coupling in the bacteriorhodopsin photocycle. Biochemistry. 1991 May 21;30(20):5016–5022. doi: 10.1021/bi00234a025. [DOI] [PubMed] [Google Scholar]
  64. Weik M., Zaccai G., Dencher N. A., Oesterhelt D., Hauss T. Structure and hydration of the M-state of the bacteriorhodopsin mutant D96N studied by neutron diffraction. J Mol Biol. 1998 Jan 30;275(4):625–634. doi: 10.1006/jmbi.1997.1488. [DOI] [PubMed] [Google Scholar]
  65. Xiao W., Brown L. S., Needleman R., Lanyi J. K., Shin Y. K. Light-induced rotation of a transmembrane alpha-helix in bacteriorhodopsin. J Mol Biol. 2000 Dec 15;304(5):715–721. doi: 10.1006/jmbi.2000.4255. [DOI] [PubMed] [Google Scholar]
  66. Zimányi L., Cao Y., Chang M., Ni B., Needleman R., Lanyi J. K. The two consecutive M substates in the photocycle of bacteriorhodopsin are affected specifically by the D85N and D96N residue replacements. Photochem Photobiol. 1992 Dec;56(6):1049–1055. doi: 10.1111/j.1751-1097.1992.tb09728.x. [DOI] [PubMed] [Google Scholar]
  67. Zscherp C., Schlesinger R., Tittor J., Oesterhelt D., Heberle J. In situ determination of transient pKa changes of internal amino acids of bacteriorhodopsin by using time-resolved attenuated total reflection Fourier-transform infrared spectroscopy. Proc Natl Acad Sci U S A. 1999 May 11;96(10):5498–5503. doi: 10.1073/pnas.96.10.5498. [DOI] [PMC free article] [PubMed] [Google Scholar]

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