FTIR Emission Spectra of Bacteriorhodopsin in a Vibrational Excited State

Abstract

The membrane protein bacteriorhodopsin (bR) con tains retinal as its chromophore. In light adapted bR, retinal is in the all trans form covalently linked to the apoprotein at Lys216 by a protonated Schiff base. Photon absorption isomerizes the retinal into the 13 cis configu ration. This primary event is responsible for both trans membrane proton transfer and subsequent re isomeriza tion of retinal (see for review [1 3]). These two processes are characterized by high quantum and thermodynamic efficiency The functioning of bR is a cyclic process that is char acterized by sequential conversion of spectrally identified intermediates (J, K, L, M, N, and O); the exact number of intermediates and the pathways of their interconver sion remains unclear. Primary light induced stages are usually described by the following chain of events Vibrational spectroscopy (resonance Raman spec troscopy) and differential Fourier transformed infrared (FTIR) spectroscopy are the most popular methods employed for the molecular study of the structure and properties of bR. However, due to high contribution of water and protein backbone to bR absorption, it is very difficult to obtain the vibrational IR spectrum of the chromophore complementary to the Raman spectrum. The use of IR emission spectroscopy would overcome these difficulties; however, we do not know of results of such studies available in the literature. Infrared emission spectroscopy has great potential. Due to the development of highly sensitive Fourier trans form infrared techniques, it is widely used in physics and chemistry In the present work, we continue these studies; we were particularly interested in the study of the effect of 0006 2979/01/6611 1315$25.00 ©2001 MAIK "Nauka / Interperiodica" * To whom correspondence should be addressed. Vol. 66, No. 11, 2001, pp. 1315 1322. Translated from Biokhimiya, Vol. 66, No. 11, 2001, pp. 1628 1637. Original Russian Text Copyright © 2001 Abstract-Vibrational IR emission spectra of bacteriorhodopsin (bR) were recorded under continuous illumination with vis ible light at room temperature. They contain selective information about the chromophore, Schiff base, and opsin. The spec tral bands were identified by comparing the data with resonance Raman and IR absorption data. The IR emission spectra were shown to contain a set of bands characteristic for both all trans (bR 568 ) and 13 cis conformations (K 610 like intermedi ate) simultaneously. Variation of spectral composition and the intensity of visible light illumination influenced the spectral traces and intensity distribution between them. Greater intensity of deformational vibrations suggests distorted retinal struc ture in the vibrationally excited ground electronic state. The origin of the emitting species of bR is discussed