We have monitored the interactions of the series of trivalent lanthanide cations with the thylakoid membrane surface of spinach chloroplasts using two complementary spectral techniques. Measurements of the fluorescence emission of the extrinsic probe 2-p-toluidinonaphthalene-6-sulfonate (TNS) and the absorbance of the intrinsic chromophore chlorophyll provide two sensitive means of characterizing the dependence of the cationmembrane interaction on the nature of the cation. In these systems, added lanthanide cations adsorb onto the membrane surface to neutralize exposed segments of membrane-embedded protein complexes. The lanthanideinduced charge neutralization increases the proximity of added TNS anion to the membrane surface as evidenced by variations in the TNS fluorescence level and wavelength of maximum emission. Our results reveal a strong dependence of TNS fluorescence parameters on both lanthanide size and total orbital angular momentum L value. Lanthanides with greater charge density (small size and/or low L value) enhance the TNS fluorescence level to a greater extent. A possible origin for the lanthanide-dependent TNS fluorescence levels is suggested in terms of a heterogeneity in the number and type of TNS binding sites. In the absence of the probe TNS, lanthanide-induced changes in the chlorophyll absorption spectrum reflect the shrinkage of chloroplasts accompanying thylakoid membrane stacking. Absorbance increases in the 500-660 nm region, attributed to increases in light scattering arising from the membrane structural reorganization, reveal a dependence on lanthanide identity. The data are consistent with the proposal that larger lanthanides with smaller enthalpies of hydration induce more significant membrane appression. These investigations illustrate a novel utilization of lanthanides in cation binding studies by employing their chemical and physical differences rather than their similarities in luminescence properties
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