Regulation of the calcium ion pump of sarcoplasmic reticulum: reversible inhibition by phospholamban and by the calmodulin binding domain of the plasma membrane calcium ion pump.
A 45 amino acid peptide (A45) corresponding to the phospholamban (PLN) binding domain of the sarcoplasmic reticulum (SR) ATPase was synthesized. Circular dichroism experiments have shown that the peptide had a predominantly random-coil conformation but adopted a higher proportion of secondary structure in the presence of a synthetic 32 amino acid peptide corresponding to the hydrophilic portion of PLN. A similar conformational change was induced by the synthetic calmodulin binding domain of the plasma membrane Ca2+ pump (peptide C28W), which acts as an endogenous inhibitor of the pump and is homologous to PLN. Cross-linking experiments have shown that peptide C28W interacted with peptide A45. The Ca(2+)-pumping activity of cardiac SR, which contains endogenous PLN, was stimulated about 30% by peptide A45. The stimulation was maximal at submicromolar Ca2+ levels and tended to disappear at higher Ca2+ concentrations. By contrast, the Ca(2+)-pumping activity of skeletal muscle SR, which lacks endogenous PLN, was unaffected. Peptide C28W strongly inhibited the pumping activity of skeletal muscle SR, and peptide A45 reversed the inhibition. The results suggest that peptide A45 competed with the ATPase for phospholamban or for peptide C28W, removing the inhibition of the pump. Thus, the exogenous inhibitor of the SR Ca(2+)-ATPase, PLN, and the internal inhibitor of the plasma membrane Ca(2+)-ATPase, peptide C28W, are functionally analogous.
Vorherr
T
T
Laboratory of Biochemistry, Swiss Federal Institute of Technology (ETH), Zurich.
Chiesi
M
M
Schwaller
R
R
Carafoli
E
E
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000187:drug effects* / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D002462:Cell Membrane / Q000737:chemistry; D002942:Circular Dichroism; D004285:Dogs; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000737:chemistry* / Q000187:drug effects / Q000201:enzymology; D012995:Solubility; D000099211:Phospholamban
D002118:Calcium; D002135:Calcium-Binding Proteins; D000252:Calcium-Transporting ATPases; D002147:Calmodulin; D010455:Peptides; D000099211:Phospholamban