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Abstract
The unicellular protozoan ciliate, Stentor coeruleus, exhibits sensitive responses to visible light. The primary photoreceptor for the photophobic and phototactic responses in this organism is a chromophore containing protein named stentorin. Two forms of the stentorin pigments, stentorin I (strongly fluorescent) and stentorin II (very weakly fluorescent), have been isolated from the pigment granules or from whole cell extracts by a hydroxylapatite and/or a Bio-Gel A-1.5m gel filtration column chromatography. These pigment preparations have been characterized by a variety of techniques, including HPLC, polyacrylamide gel electrophoresis and spectroscopy. The apparent molecular weights of stentorin I and II have been determined to be 82,000-103,000 and 500,000-810,000, respectively. Stentorin I is regarded as a proteolipid which shows anomalous behavior on SDS-PAGE and seems to form oligomeric complexes in solution, and is either a non-functional fraction or serves as an antenna pigment. Stentorin II appears to be the primary photoreceptor whose absorption and fluorescence properties are consistent with the action spectra for the photoresponses of the ciliate to visible light. Stentorin II exhibits typical properties of a membrane protein. The chromophore in stentorin II has an induced optical activity, most likely due to a protein-chromophore interaction, that is strong enough to withstand mild detergent conditions. This may indicate that the protein-chromophore interaction is of a covalent character. Stentorin II exhibits four fluorescence decay components at various emission wavelengths, pHs and pDs. Investigation of the effects of deuterium oxide and pD, as well as pH, on the fluorescence decay kinetics and time-resolved spectra of stentorins shows that stentorin I exhibits a free chromophore-like peptide behavior, as judged by its accessibility to solvents, whereas stentorin II is a larger molecular assembly composed of several proteins in which the chromophore may be deeply imbedded within the hydrophobic core of the protein. These results are consistent with a model showing proton dissociation as a primary photoprocess in the excited state of stentorin, facilitated by the quarternary structure and/or integrity of the protein assembly of the photoreceptor structure.
