title: Mechanism of action of the novel plasma membrane Ca(2+)-pump inhibitor caloxin.
abstract: Caloxin 2A1 is a novel inhibitor of the plasma membrane (PM) Ca(2+)-pump [Am. J. Physiol. Cell Physiol. 280 (2001) C1027]. The PM Ca(2+)-pump is a Ca(2+)-Mg(2+)-ATPase that expels Ca(2+) from cells to help them maintain low concentrations of cytosolic Ca(2+). Caloxin 2A1 inhibits Ca(2+)-Mg(2+)-ATPase in human erythrocyte leaky ghosts. Here we report that this inhibition is non-competitive with respect to the substrates Ca(2+) and ATP and the activator calmodulin. This was anticipated since the high affinity binding site for Ca(2+) and sites for ATP and calmodulin are intracellular whereas caloxin 2A1 is a peptide selected for binding to the second extracellular domain of the pump. Caloxin 2A1 also inhibited the Ca(2+)-dependent formation of the acid stable 140 kDa acylphosphate intermediate from 32P-gamma-ATP. However, it did not inhibit the formation of the acylphosphate intermediate in the reverse direction-from 32P-orthophosphate. Consistent with results on mutagenesis of transmembrane residues in the pump protein, we suggest that caloxin 2A1 inhibits conformational changes required during the reaction cycle of the pump.
authors.affiliation: Department of Biology, McMaster University, Hamilton, Ont, Canada L8N 3Z5.
authors.forename: M E
authors.initials: ME
authors.lastname: Holmes
authors.forename: J
authors.initials: J
authors.lastname: Chaudhary
authors.forename: A K
authors.initials: AK
authors.lastname: Grover
mesh_terms: D017766:Acid Anhydride Hydrolases / Q000187:drug effects / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites / Q000187:drug effects / Q000502:physiology; D001667:Binding, Competitive / Q000187:drug effects / Q000502:physiology; D017301:Ca(2+) Mg(2+)-ATPase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects* / Q000502:physiology; D002147:Calmodulin / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004912:Erythrocytes; D005057:Eukaryotic Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides / Q000494:pharmacology*; D011487:Protein Conformation / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000187:drug effects / Q000502:physiology; D013347:Subcellular Fractions; D000097024:Acylphosphatase
chemical_list: D017766:Acid Anhydride Hydrolases; D000255:Adenosine Triphosphate; D017301:Ca(2+) Mg(2+)-ATPase; D002118:Calcium; D002147:Calmodulin; D004791:Enzyme Inhibitors; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides; D000097024:Acylphosphatase; C423122:caloxin 2A1