Measurement of transmembrane potentials in Rhodospirillum rubrum chromatophores with an oxacarbocyanine dye.
The fluorescent dye 3,3-dipentyloxacarbocyanine (OCC) can be used as a fluorescence probe to measure transmembrane potentials across Rhodospirillum rubrum chromatophore membranes. A reversible fluorescence increase is observed in the light which is sensitive to inhibitors, permeable ions and uncouplers. Partial interchangeability between the electrical potential and the proton concentration gradient has been demonstrated by measurement of the fluorescence increase with OCC and the fluorescence quenching with 9-aminoacridine. OCC fluorescence changes can be induced also in the dark by injection of permeable salts and by rapid pH changes presumably indicating diffusion potentials. Using salt-induced diffusion potentials for calibrating the light signals and with several assumptions, the light-induced potentials were estimated as 170 mV for the maximal signal and 90-110 mV at the steady state. OCC has been shown to apparently increase the electrical conductivity of the chromatophore membrane, a fact which may be relevant to the mechanism of action of this probe. A red shift in the OCC absorption spectrum occurs when mixed with chromatophores, with a difference spectrum maximum at 495 nm. The absorption changes at 495 nm taking place in the light are similar in kinetics to the fluorescence changes. The absorbance spectrum of OCC in organic solvents is red shifted and the extent of the shift depends on the hydrophobicity of the medium. The difference spectrum compared to water in sec-butyl acetate/n-hexane (3 : 1, v/v) with a dipole moment of 5 was nearly identical to that of chromatophore-associated dye. The uncoupling properties of OCC at high concentrations and some difficulties in calibration limit the usefulness of this probe for quantitative measurements of transmembrane potentials.
D020130:Bacterial Chromatophores / Q000187:drug effects / Q000378:metabolism*; D002232:Carbocyanines; D002794:Choline / Q000378:metabolism; D005456:Fluorescent Dyes; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects; D011189:Potassium Chloride / Q000494:pharmacology; D011804:Quinolines; D012247:Rhodospirillum rubrum / Q000378:metabolism*; D012492:Salts / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D014634:Valinomycin / Q000494:pharmacology
D002232:Carbocyanines; D005456:Fluorescent Dyes; D011804:Quinolines; D012492:Salts; D014634:Valinomycin; D011189:Potassium Chloride; D002794:Choline