Flash photolysis study on pharaonis phoborhodopsin from a haloalkaliphilic bacterium (Natronobacterium pharaonis)

• DOI: 10.1016/0005-2728(92)90015-t
• Publication Date: 2003-02-10T23:57:53Z
• Authors: Mamoru Miyazaki; Junichi Hirayama; Masaki Hayakawa; Naoki Kamo
• Keywords: 0301 basic medicine; 03 medical and health sciences

Abstract Pharaonis phoborhodopsin (ppR) functions as a photoreceptor of the negative phototaxis of haloalkaliphilic bacterium (Natronobacterium pharaonis). The absorbance maximum of ppR is located at 498 nm. ppR has a cyclic photo-reaction (photo cycle): upon illumination, ppR transforms to a photo-intermediate which returns to the original pigment through various intermediates in the dark reaction. By flash-photolysis in a millisecond time scale, the photocycle of ppR was investigated. Two photointermediates were found whose λmax was 390 nm and 550 nm. The former is referred to as ppRM, and the latter as ppRO since these correspond respectively to M- and O-intermediates of bacteriorhodopsin. Flash photolysis data were kinetically analyzed according to the cyclic sequential scheme of ppR → ppRM → ppRO → ppR. In the analysis of kinetic data of one wavelength representing the change in an intermediate, the contribution from other intermediates or the original pigment was considered because their absorption maxima are close to each other. The rate constant of ppRM → ppRO was taken as k1, and that of ppRO → ppR, k2. k1 depended greatly on pH in the medium, while k2 had little dependence on pH. The activation energies of k1 and k2 were determined. Addition of azide (100 mM, pH 7, 20°C) led to about 40-fold increase of k1 while the influence on k2 was slight.