Bacteriorhodopsin protonation pathways

Different aspects of bacteriorhodopsin functioning have been studied early [35-37, 223, 233-249]. However, as a rule, the researchers have described the mechanism of proton transport phenomenologically, though the analysis of the results taken from the FT-IR difference spectra yielded detailed descriptions of the proton state in a proton pathway in all of the above-mentioned intermediates (see, e.g., Zundel [6]). 

B. Brzezinski, P. Radziejewski, J. Olejnik and G. Zundel, An intramolecular hydrogen-bonded system with large proton polarizability - A model with regard to the proton pathway in bacteriorhodopsin and other systems with collective proton motion, J. Mol. Struct., 323 (1994) 71. 

Fig. 2. A tentative scheme of the bacteriorhodopsin pump. bR indicates the bacteriorhodopsin ground state, and L, M, N(P) and O indicate the corresponding intermediates of the photocycle. NHs, =N2 and =NH represent the protonated Schiff base of the idUtrans retinal residue, the deprotonated and the protonated Schiff bases of 13-cw retinal residues, respectively. -COOH and -COO are the protonated and the deprotonated Asp-96 carboxylic group, respectively. The outward hydrophilic H -conducting pathway (the proton well) is shaded. (From Skulachev[35].)...

Fig. 5. Suggested scheme for the photocycle of bacteriorhodopsin [114-116], The photoreaction produces state J, the other reactions represent the pathway of thermal relaxation back to the initial state. Where release and uptake are shown, they refer to exchange of the proton with the aqueous...

The 13-c/j retinal-chromophore in dark-adapted bacteriorhodopsin exhibits a very different photocycle, whose predominant intermediate has an absorption maximum at 610 nm [199], and which contains no intermediate [202,238] analogous to M. The 610 nm intermediate will decay to either the 13-c/s chromophore or the dW-trans form, the latter pathway being responsible for the phenomenon of light-adaptation [199]. This pathway does not explain, however, why monomeric bacteriorhodopsin shows poor light-adaptation [168,239]. The chromophore in the 13-c/s configuration is not associated with proton translocation [240]. Indeed, reconstitution of bacterio-opsin with 13-demethyl retinal, which traps the retinal moiety in the 13-c/s configuration, results [241] in a non-transporting photocycle. 

Theoretical models for the functioning of bacteriorhodopsin must include entry and leaving sites for protons on the two sides of the membrane, a proton conduction pathway, and the unidirectional translocation of protons across a potential barrier somewhere inside the protein so as to accomplish net transport against an electro-... 

Proton transfer is closely linked to the structure of the reaction-center protein. Since protons are present in the external aqueous medium, the (reduced) quinone molecules are buried inside the interior ofthe reaction-center protein, therefore protonation would seem to require some kind of channel for the passage of water molecules. However, at least until recently (see below), there was no evidence for the presence of channels large enough to accommodate water molecules. An alternative mechanism might involve a chain of ionizable amino acids which extends from the surface of the protein to the interior where the reduced quinone is located, forming a pathway along which protons may be transported. Such a mechanism has been likened to a bucket brigade or relay station and shown to exist in such proteins as bacteriorhodopsin, ATP synthase and cytochrome oxidase. 

HPLC analysis also revealed that the protonated Schiff base of all-traws-retinal in solution is isomerized predominantly into the 11-cis form (82% 11-cis, 12% 9-cis, and 6% 13-ds in methanol) [23]. The 11-cis form as a photoproduct is the nature of retinochrome, not those of archaeal rhodopsins. This suggests that the protein environment of retinochrome serves as the intrinsic property of the photoisomerization of the retinal chromophore. In contrast, it seems that the protein environment of archaeal rhodopsins forces the reaction pathway of the isomerization to change into the 13-cis form. In this regard, it is interesting that the quantum yield of bacteriorhodopsin (0.64) is 4—5 times higher than that in solution (-0.15) [21,23], The altered excited state reaction pathways in archaeal rhodopsins never reduce the efficiency. Rather, archaeal rhodopsins discover the reaction pathway from the all-trans to 13-cis form efficiently. Consequently, the system of efficient isomerization reaction is achieved as well as in visual rhodopsins. Structural and spectroscopic studies on archaeal rhodopsins are also reviewed in Section 4.3. 

The experimental structure of bR determined at atomic resolution from cryoelectron microscopy and X-ray crystallography revealed a channel containing the Schiff base of the retinal chromophore (27, 28). Site-directed mutagenesis and vibrational spectroscopy experiments have enabled the identification of polar residues in the channel involved in the proton transfer pathway (29-32). Recent work on bacteriorhodopsin has concentrated on hydration and conformational thermodynamics. 

Such a mechanism, analogous to a bucket bri- /r/ 5 Backbone of the L subunit (heavy lines), M subunit gade", has been proposed for proton transfer (light lines) and cofactors. The van der Waals surface of across biological membranes (16) and has been (dotted) outline a possible pathway for Qg to leave the supported by observations on bacteriorhodopsin RC. Modified from ref. 12. 

H. Luecke, H.T. Richter, J.K. Lanyi, Proton transfer pathways in bacteriorhodopsin at 2.3 angstrom resolution. Science 280 (1998) 1934—1937. 

I. Kawamura, M. Ohmine, J. Tanabe, S. Tuzi, H. Saito, A. Naito, Dynamic aspects of extracellular loop region as a proton release pathway of bacteriorhodopsin studied by relaxation time measurements by solid state NMR, Biochim. Biophys. Acta 1768 (2007) 3090-3097. 

See, N.A. Dencher, G. Boldt, J. Heberle, H.-D. Hdltje and M. Hditje, in T. Bounds (Ed.), Proton-Transfer in Hydrogen-Bonded Systems, NATO ASI, Ser. B, Vol. 291, Plenum Press, 1992, pp. 171-186. On p. 180 of this paper there is a diagram of a computer-graphics model of the pathways for protons across bacteriorhodopsin. 

Rammelsberg, R., Huhn, G., Luebben, M., and Gerwert, K., Bacteriorhodopsin s intramolecular proton-release pathway consists of a hydrogen-bonded network. Biochemistry, 37, 14, 5001-5009, 1998. 

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