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Spectroscopy
Volume 16 (2002), Issue 3-4, Pages 107-120
http://dx.doi.org/10.1155/2002/190968

Dynamic aspect of bacteriorhodopsin as viewed from 13C NMR: Conformational elucidation, surface dynamics and information transfer from the surface to inner residues

H. Saitô, R. Kawaminami, M. Tanio, T. Arakawa, S. Yamaguchi, and S. Tuzi

Department of Life Science, Himeji Institute of Technology, Harima Science Garden City, Kamigori, Hyogo 678-1297, Japan

Copyright © 2002 Hindawi Publishing Corporation. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

We demonstrate here how dynamic as well as conformational features of bacteriorhodopsin (bR) in purple membrane (PM) as a typical membrane protein are revealed by extensive 13C NMR studies on [3-13C]-, [2-13C]-, [1-13C]Ala or [1-13C]Val-labeled bR and a variety of site-directed mutants. A number of 13C NMR peaks were well-resolved for [3‒13C]Ala‒ and [1-13C]Val-bR under the condition of fully hydrated PM at ambient temperature and assigned to individual amino-acid residues, initially by regio-specific manner with reference to the data of the conformation-dependent displacements of peaks from model polypeptides and subsequently by site-specific manner with reference to the specifically reduced peak-intensities of site-directed mutant as compared with those of wild type. It is noticeable that the revealed bR structure at ambient temperature by 13C NMR is not static as anticipated from the data of diffraction studies at cryo-temperature but is dynamically heterogeneous undergoing motional fluctuations with various frequencies (102–108 Hz) depending upon the domains of interest. We further applied this approach to reveal how charged state of surface residues, especially at the side-chain of exracellular Glu residues (Glu 194 and 204), could be transmitted to the inner part of the helices such as Ala 53, 84, and 215 to alter their local conformations of transmembrane helices near at the Schiff base through side-chain interactions. We also analyzed how information of the protonation at Asp 85 from helix C is initially transmitted to helices B (Val 49) and G (Val 213) though modified helix‒helix interactions through the side-chains of Arg 82.