Proton release and the photocycle of bacteriorhodopsin

Bacteriorhodopsin (bR) is a light-driven proton pump. Over the last several years, much has been done in determining the structure and function of this protein; however, the exact mechanism of the proton pump is still unknown. My work presented here focuses on characterizing the proton release process of bR. By studying the pH dependence, mutations of Arg-82 and Tyr-57, modifications of surface charges, and comparisons with two archaerhodopsins, I suggest that the proton release group has an intrinsic pK of 8.2 and is likely Arg-82 or groups interacting with Arg-82. Proton release is inhibited at high pH. The alkaline form of bR is also characterized by a faster formation of the M intermediate and a large photocurrent component. These features are present in the R82A mutant, which has greatly delayed proton release. Earlier suggestions of a tyrosine deprotonation being requisite for the alkaline form of bR was not confirmed in this study. Tyr-57, however, does affect the kinetics and efficiency of proton release. Of the charged groups on the surface of the membrane, only Lys-129 appears to greatly inhibit normal light-driven proton release. The negatively charge lipids and carboxylates do not appear to regulate proton release; however, when surface lysines are modified, the efficiency of normal proton release is reduced. In support of this are proton release measurements from two archaerhodopsins (aR-1 and aR-2) which are proton pumps similar to bR. Coupled with published results, my studies suggest that Arg-82 is very important in light-driven proton release. Its interaction with Asp-85 maintains a favorable interaction between Asp-85 and the protonated Schiff base. Mutations in this region of bR can alter the kinetics of the photocycle and proton release by disrupting the native orientations and distances between these residues. On the surface of the protein, Lys-129 appears to facilitate the final release of protons into the external medium.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biophysics, General

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2011

Author dc:creator

• Kono, Masahiro

Contributors dc:contributor

• Ebrey, Thomas G.

• Biophysics, General

Statement dc:rights

• Copyright 1994 Kono, Masahiro

Language dc:language

eng

Identifier

AAI9512434
(UMI)AAI9512434

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/23779