Molecular mechanism of regulation of Ca2+ pump ATPase by phospholamban in cardiac sarcoplasmic reticulum. Effects of synthetic phospholamban peptides on Ca2+ pump ATPase.
The molecular mechanism of the regulation of Ca2+ pump ATPase by phospholamban in cardiac sarcoplasmic reticulum was examined using synthetic peptides of phospholamban and purified Ca2+ pump ATPase from cardiac sarcoplasmic reticulum. The phospholamban monomer of 52 amino acid residues contains two distinct domains, the cytoplasmic (amino acids 1-30) and the transmembrane (amino acids 31-52) domains. The peptide corresponding to the amino acids 1-31 of phospholamban (PLN 1-31) decreased the Vmax of the Ca(2+)-dependent ATPase activity in dose-dependent manner, while it had no effect on the affinity of the ATPase for Ca2+ (KCa). On the other hand, the peptide corresponding to the amino acids 28-47 of phospholamban (PLN 28-47) increased the KCa from 0.52 to 1.33 microM without significant change in the Vmax value when reconstituted into vesicles with the ATPase. Essentially the same results as PLN 28-47 were obtained with the peptide corresponding to the amino acids 8-47 of phospholamban (PLN 8-47). The inhibitory effects of PLN 1-31 and PLN 8-47 on the ATPase were reversed by cAMP-dependent phosphorylation of the peptides (Ser16). These results indicate that phospholamban suppresses Ca2+ pump ATPase at two different sites, the cytoplasmic domain for Vmax and the transmembrane domain for KCa, and that cAMP-dependent phosphorylation de-suppresses these inhibitory effects on the ATPase.
Sasaki
T
T
First Department of Medicine, Osaka University School of Medicine, Japan.
Inui
M
M
Kimura
Y
Y
Kuzuya
T
T
Tada
M
M
D000595:Amino Acid Sequence; D000818:Animals; D002135:Calcium-Binding Proteins / Q000494:pharmacology*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D004285:Dogs; D006352:Heart Ventricles; D007700:Kinetics; D008081:Liposomes; D009206:Myocardium / Q000201:enzymology*; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000494:pharmacology*; D010766:Phosphorylation; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D000099211:Phospholamban
D002135:Calcium-Binding Proteins; D000252:Calcium-Transporting ATPases; D008081:Liposomes; D010446:Peptide Fragments; D000099211:Phospholamban