Open AccessCharacterization of ATP‐dependent Ca 2+ uptake by canine brain microsomes with saponin
Author(s) -
INAMITSU Tetsuaki,
OHTSUKI Iwao
Publication year - 1984
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1984.tb08529.x
Subject(s) - endoplasmic reticulum , saponin , vesicle , microsome , membrane , chemistry , biochemistry , calmodulin , biophysics , biology , in vitro , enzyme , medicine , alternative medicine , pathology
ATP‐dependent Ca 2+ uptake by brain microsomes was classified into two fractions according to the sensitivity to saponin. Properties of each fraction of Ca 2+ uptake were examined and compared with those of inside‐out membrane vesicles of erythrocyte and cardiac sarcoplasmic reticulum. The concentration of saponin for 50 % inhibition (IC50) of major saponin‐sensitive Ca 2+ uptake was 11 μg/ml, and this uptake was enhanced by calmodulin. The minor saponin‐insensitive Ca 2+ uptake fraction (IC50;90 μg/ml) was not affected by calmodulin but was enhanced by oxalate or 0.1 M KC1. The IC50 of saponin for inside‐out membrane vesicles of erythrocyte and cardiac sarcoplasmic reticulum was 11.3 and 114.8 μg/ml, respectively. A characteristic ring‐like saponin‐cholesterol micellar structure was observed electron microscopically in most membrane vesicles of brain microsomes and erythrocyte membrane vesicles but not in the cardiac sarcoplasmic reticulum. These observations indicate that saponin‐sensitive and insensitive Ca 2+ uptake was derived from plasma membranes and endoplasmic reticulum, respectively. Saponin proved useful for distinguishing the Ca 2+ transport activity of plasma membrane from the Ca 2+ uptake of other cellular organelles in the membrane preparations.