COMPARISON OF THE KINETIC EFFECTS OF PHOSPHOLAMBAN PHOSPHORYLATION AND ANTI-PHOSPHOLAMBAN MONOCLONAL-ANTIBODY ON THE CALCIUM-PUMP IN PURIFIED CARDIAC SARCOPLASMIC-RETICULUM MEMBRANES
Ay. Antipenko et al., COMPARISON OF THE KINETIC EFFECTS OF PHOSPHOLAMBAN PHOSPHORYLATION AND ANTI-PHOSPHOLAMBAN MONOCLONAL-ANTIBODY ON THE CALCIUM-PUMP IN PURIFIED CARDIAC SARCOPLASMIC-RETICULUM MEMBRANES, Biochemistry, 36(42), 1997, pp. 12903-12910
Protein kinase A-(PKA-) catalyzed phosphorylation of phospholamban (PL
N), the protein regulator of the cardiac Ca pump, mediates abbreviatio
n of systole in response to beta-adrenergic agonists. Investigators pr
eviously, however, have been unsuccessful in demonstrating an effect o
f PLN phosphorylation or anti-PLN monoclonal antibody (mAb), which is
considered to mimic phosphorylation's well-known effect on K-m(Ca), on
microsomal Ca uptake at the (high) Ca2+ concentrations found intracel
lularly at peak systole. We therefore compared the effects of the cata
lytic subunit of PKA and anti-PLN mAb on the kinetics of Ca uptake in
sucrose gradient-purified cardiac microsomes. Both treatments produced
a 33-44% increase in V-max(Ca) at 25 and 37 degrees C, and an 11-31%
decrease in K-m(Ca), with comparable changes in Ca2+-ATPase activity.
An acceleration of E2P decomposition upon PLN phosphorylation may cont
ribute to the increased V-max(Ca) of Ca uptake at 25 degrees C but not
at 37 degrees C, based on measurement of the kinetics of E2P decompos
ition and steady-state E2P formation from P-i at different temperature
s. Our data document almost identical increases in V-max(Ca) of micros
omal Ca uptake with PLN phosphorylation or addition of anti-PLN mAb an
d hence provide insight into the kinetic mechanism of PLN's regulation
of the cardiac sarcoplasmic reticulum Ca pump protein.