A quench-flow kinetic investigation of calcium ion accumulation by isolated cardiac sarcoplasmic reticulum. Dependence of initial velocity on free calcium ion concentration and influence of preincubation with a protein kinase, MgATP, and cyclic AMP.

Ca2+ accumulation at pH 6.8 by isolated rabbit heart microsomes derived chiefly from sarcoplasmic reticulum was investigated by a quench-flow technique. The reaction was terminated at preset times by addition to the reaction mixture of an equal volume of 10 to 50 mM ethyleneglycol-bis-(beta-aminoethyl ether)-N,N'-tetraacetic acid buffered at pH 6.0. The initial velocity of Ca2+ accumulation by microsomal preparations exhibiting a steady state Ca2+ accumulation of 25.6 nmol Ca2+/mg increased from 3.67 to 33.4 nmol Ca2+/mg - s as the free Ca2+ concentration was raised from 0.2 to 18.9 muM. Preincubation of the cardiac microsomes with a partly purified soluble cardiac cyclic AMP-dependent protein kinase, MgATP, and cyclic AMP lead to a significant increase in the initial Ca2+ accumulation rate. The amounts of Ca2+ that were found to accumulate in the first 200 ms of the reaction are comparable to the quantities of the ion that according to literature data need to be removed from the myofilaments and the myoplasm for induction of relaxation of the myocardial fibers.