Effects of eosin Y on the catalytic and functional activities of Mg2+,ATP-dependent calcium pump of smooth muscle cell plasma membrane.
Experiments with highly purified preparations of Ca2+,Mg2+-ATPase solubilized from sarcolemma and a fraction of inside-out sarcolemmal vesicles were performed to study the kinetics of inhibitory effects of eosin Y (0-100 microM) on the catalytic and transport activity of Mg2+,ATP-dependent calcium pump of myometrial cell plasma membrane. For both the Ca2+,Mg2+-dependent ATP hydrolysis and the Mg2+, ATP-dependent accumulation of Ca2+ the apparent inhibitory constant Ki was 0.8 microM. However, eosin Y used at concentrations of up to 100 microM had absolutely no effect on Na+-Ca2+ exchange in the plasma membrane fraction. This inhibitor decreased the turnover rate of purified Ca2+,Mg2+-ATPase determined both by Mg2+ and ATP. However, the affinity of purified Ca2+,Mg2+-ATPase for Mg2+ increased, and its affinity for ATP decreased in the presence of eosin Y. In the case of Mg2+,ATP-dependent Ca2+ transport, eosin Y decreased the turnover rate of the calcium pump whose affinity for Mg2+ and ATP displayed virtually no dependence on the presence of the inhibitor in the incubation medium. The possibility of the use of eosin Y in model experiments for identification of Mg2+, ATP-dependent and Na+-dependent calcium fluxes from smooth muscle cells into the intercellular milieu is discussed.
Department of Muscle Biochemistry, Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kiev, 252030, Ukraine.
N N
NN
Slinchenko
N F
NF
Bratkova
S A
SA
Kosterin
V P
VP
Zimina
I G
IG
Chernysh
D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism*; D002384:Catalysis; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D004801:Eosine Yellowish-(YS) / Q000494:pharmacology*; D007700:Kinetics; D009130:Muscle, Smooth / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D013552:Swine
D017301:Ca(2+) Mg(2+)-ATPase; D004801:Eosine Yellowish-(YS)