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1 May 2001 Trapping and Spectroscopic Identification of the Photointermediates of Bacteriorhodopsin at Low Temperatures
Sergei P. Balashov, Thomas G. Ebrey
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Light-driven transmembrane proton pumping by bacteriorhodopsin occurs in the photochemical cycle, which includes a number of spectroscopically identifiable intermediates. The development of methods to crystallize bacteriorhodopsin have allowed it to be studied with high-resolution X-ray diffraction, opening the possibility to advance substantially our knowledge of the structure and mechanism of this light-driven proton pump. A key step is to obtain the structures of the intermediate states formed during the photocycle of bacteriorhodopsin. One difficulty in these studies is how to trap selectively the intermediates at low temperatures and determine quantitatively their amounts in a photosteady state. In this paper we review the procedures for trapping the K, L, M and N intermediates of the bacteriorhodopsin photocycle and describe the difference absorption spectra accompanying the transformation of the all-trans-bacteriorhodopsin into each intermediate. This provides the means for quantitative analysis of the light-induced mixtures of different intermediates produced by illumination of the pigment at low temperatures.

Sergei P. Balashov and Thomas G. Ebrey "Trapping and Spectroscopic Identification of the Photointermediates of Bacteriorhodopsin at Low Temperatures," Photochemistry and Photobiology 73(5), 453-462, (1 May 2001).<0453:TASIOT>2.0.CO;2
Received: 11 December 2000; Accepted: 1 February 2001; Published: 1 May 2001

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