Effect of spermine on the activity of synaptosomal plasma membrane Ca(2+)-ATPase reconstituted in neutral or acidic phospholipids.
The activity of purified plasma membrane Ca(2+)-ATPase (PMCA) from pig brain was inhibited by spermine (a naturally occurring and highly abundant polycation in brain). The level of inhibition was dependent on the phospholipid used for reconstitution as well as on the intact or truncated state of the enzyme. An IC(50) value of 12.5 mM spermine was obtained for both, the intact protein plus calmodulin and the trypsin-digested protein, reconstituted in phosphatidylcholine (PC). In the absence of calmodulin the intact Ca(2+)-ATPase gave an IC(50) of 27 mM. This form was more sensitive to spermine inhibition when it was reconstituted with phosphatidylserine (PS), showing an IC(50) value of 2.5 mM spermine. However, the truncated form was less responsive to spermine inhibition, having an IC(50) value of 12.5 mM. Spermine has no effect on the affinity of the PMCA for Ca(2+) or ATP, but its effect on the protein is pH-dependent. It is suggested that spermine could bind to negatively charged residues on the ATPase with different accessibility, depending on the structural rearrangement of the protein. Further, when the protein is reconstituted in PS, spermine also binds to the lipid.
Palacios
Javier
J
Departamento de Bioquímica y Biología Molecular y Genética, Facultad de Ciencias, Universidad de Extremadura, 06071 Badajoz, Spain.
Sepúlveda
M Rosario
MR
Salvador
J M
JM
Mata
Ana M
AM
D000255:Adenosine Triphosphate; D000818:Animals; D002118:Calcium; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors*; D002147:Calmodulin / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D010743:Phospholipids; D013096:Spermine / Q000494:pharmacology*; D013552:Swine; D013574:Synaptosomes / Q000187:drug effects* / Q000201:enzymology; D014357:Trypsin
D002147:Calmodulin; D004791:Enzyme Inhibitors; D010743:Phospholipids; D013096:Spermine; D000255:Adenosine Triphosphate; D014357:Trypsin; D000252:Calcium-Transporting ATPases; D002118:Calcium