The effects of membrane potential and lanthanides on the conformation of the Ca2+-transport ATPase in sarcoplasmic reticulum.
The effects of Ca2+, lanthanide ions (Gd3+, La3+ and Pr3+) and membrane potential on the fluorescence of tryptophan and covalently bound fluorescein were analysed in native and fluorescein isothiocyanate (FITC)-labelled sarcoplasmic reticulum vesicles. The binding of Ca2+ and lanthanides to the Ca2+-ATPase increases the fluorescence intensity of tryptophan and decreases the fluorescence intensity of FITC; the dependence of these effects on cation concentration is consistent with the involvement of the high-affinity Ca2+-binding sites of the Ca2+-ATPase in the cation-induced fluorescence changes. The fluorescence of FITC-labelled sarcoplasmic reticulum vesicles is also influenced by membrane potential changes induced by ion substitution. Inside positive potential increases, while inside negative potential decreases, the fluorescence of bound FITC. Smaller potential-dependent changes in tryptophan fluorescence were also observed. The effects of Ca2+, lanthanides and membrane potential on the fluorescence of tryptophan and FITC are discussed in terms of the two major conformations of the Ca2+-ATPase (E1 and E2), that are assumed to alternate during Ca2+ transport. The observations support the suggestion [Dux, Taylor, Ting-Beall & Martonosi (1985) J. Biol. Chem. 260, 11730-11743] that the vanadate-induced crystals of Ca2+-ATPase represent the E2, while the Ca2+ and lanthanide-induced crystals the E1, conformation of the enzyme.
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D004533:Egtazic Acid / Q000494:pharmacology; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D008564:Membrane Potentials; D008674:Metals, Rare Earth / Q000494:pharmacology*; D008861:Microsomes / Q000378:metabolism; D011487:Protein Conformation / Q000187:drug effects; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000201:enzymology*; D013050:Spectrometry, Fluorescence; D013861:Thiocyanates; D014364:Tryptophan / Q000378:metabolism
D005452:Fluoresceins; D008674:Metals, Rare Earth; D013861:Thiocyanates; D004533:Egtazic Acid; D014364:Tryptophan; D000252:Calcium-Transporting ATPases; D016650:Fluorescein-5-isothiocyanate; D002118:Calcium