Illumination-Dependent Changes in the Intrinsic Fluorescence of Bacteriorhodopsin

Roberto A. Bogomolni, Lisa Stubbs, Janos K. Lanyi

Research output: Contribution to journalArticlepeer-review

Abstract

Bacteriorhodopsin fluorescence in the ultraviolet region shows a broad band around 330 nm. Actinic illumination in the visible band (4 μs flash, 575 nm) elicits cyclic fluorescence and absorbance changes in the ultraviolet. The fluorescence decreases with a half-time of about 40 μs at 22 °C and 120 μs at 12 °C closely following the absorbance change at 410 nm due to photointermediate M. The recovery time, however, is longer than the absorbance change at 410 nm and corresponds to that of the regeneration of bacteriorhodopsin. The fractional changes in fluorescence span a broad spectral range with peaks at about 330, 350, and 385 nm. Absorbance changes in the ultraviolet show an increase between 330 and 400 nm corresponding to the tail of the 410-nm peak, and a spectral shift to longer wavelength with maximum and minimum at 300 and 275-285 nm, respectively. The fluorescence emission spectrum at 1 ms after the flash (corresponding largely to the photointermediate M410) was calculated and shows peaks at 304 and 328 nm and a broad shoulder at longer wavelengths, having 90%, 50%, and 35% (or less) of the original bacteriorhodopsin fluorescence intensity, respectively. Cesium chloride (1 M) selectively quenches the 350-nm fluorescence and markedly decreases the fluorescence change in that region. These results suggest that of the tryptophans in bacteriorhodopsin one or more is exposed to a polar environment (for example, the aqueous medium), and such tryptophan(s) as well as others are significantly quenched in the transition to M410. The fluorescence changes may reflect changes in the spatial disposition of these residues, in their interaction with the chromophore, or in their ionization state.

Original languageEnglish (US)
Pages (from-to)1037-1041
Number of pages5
JournalBiochemistry
Volume17
Issue number6
DOIs
StatePublished - Jan 1 1978
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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