title: The effects of calcium, temperature and phospholamban phosphorylation on the dynamics of the calcium-stimulated ATPase of canine cardiac sarcoplasmic reticulum.
abstract: Highly purified sarcoplasmic reticulum (SR) has been prepared from dog hearts and has been incubated with the triplet probe erythrosinyl isothiocyanate to specifically label the Ca2+-stimulated ATPase (Ca2+-ATPase) of the SR. The rotational mobility of the Ca2+-ATPase has been studied in this erythrosin-labelled SR using time-resolved phosphorescence polarization. Qualitatively, the mobility of the cardiac Ca2+-ATPase resembles that of skeletal muscle SR Ca2+-ATPase. Addition of Ca2+ to SR affects the mobility of the Ca2+-ATPase in a way consistent with a segment of the ATPase altering its orientation relative to the plane of the membrane. Phosphorylation of phospholamban in cardiac SR by the purified catalytic subunit of cAMP-dependent protein kinase, which is known to increase the activity of the Ca2+-ATPase by deinhibition, also alters measured anisotropy. The changes observed are not compatible with dissociation of the Ca2+-ATPase from phospholamban after the latter is phosphorylated. The data are more consistent with phospholamban associating with the Ca2+-ATPase following phosphorylation, or more complex models in which only the hydrophilic domain of phospholamban binds with and dissociates from the Ca2+-ATPase.
authors.affiliation: Department of Protein and Molecular Biology, Royal Free Hospital School of Medicine, London, U.K.
authors.forename: C
authors.initials: C
authors.lastname: Fowler
authors.forename: J P
authors.initials: JP
authors.lastname: Huggins
authors.forename: C
authors.initials: C
authors.lastname: Hall
authors.forename: C J
authors.initials: CJ
authors.lastname: Restall
authors.forename: D
authors.initials: D
authors.lastname: Chapman
mesh_terms: D000818:Animals; D002118:Calcium / Q000502:physiology*; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002458:Cell Fractionation / Q000379:methods; D002499:Centrifugation, Density Gradient; D004285:Dogs; D004923:Erythrosine; D005260:Female; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D007700:Kinetics; D008297:Male; D009206:Myocardium / Q000201:enzymology*; D010766:Phosphorylation; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013696:Temperature; D013861:Thiocyanates; D000099211:Phospholamban
chemical_list: D002118:Calcium; D002135:Calcium-Binding Proteins; D000252:Calcium-Transporting ATPases; D004923:Erythrosine; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D013861:Thiocyanates; D000099211:Phospholamban