A high affinity acceptor for phospholipase A2 with neurotoxic activity is a calmodulin.
One of the high affinity binding proteins for ammodytoxin C, a snake venom presynaptically neurotoxic phospholipase A(2), has been purified from porcine cerebral cortex and characterized. After extraction from the membranes, the toxin-binding protein was isolated in a homogenous form using wheat germ lectin-Sepharose, Q-Sepharose, and ammodytoxin-CH-Sepharose chromatography. The specific binding of (125)I-ammodytoxin C to the isolated acceptor was inhibited to different extents by some neurotoxic phospholipases A(2), ammodytoxins, bee venom phospholipase A(2), agkistrodotoxin, and crotoxin; but not by nontoxic phospholipases A(2), ammodytin I(2), porcine pancreatic phospholipase A(2), and human type IIA phospholipase A(2); suggesting the significance of the acceptor in the mechanism of phospholipase A(2) neurotoxicity. The isolated acceptor was identified as calmodulin by tandem mass spectrometry. Since calmodulin is generally considered as an intracellular protein, the identity of this acceptor supports the view that secretory phospholipase A(2) neurotoxins have to be internalized to exert their toxic effect. Moreover, since ammodytoxin is known to block synaptic transmission, its interaction with calmodulin as an acceptor may constitute a valuable probe for further investigation of the role of the latter in this Ca(2+)-regulated process.
Department of Biochemistry and Molecular Biology, Jozef Stefan Institute, Jamova 39, Slovenia.
J
J
Sribar
A
A
Copic
A
A
Paris
N E
NE
Sherman
F
F
Gubensek
J W
JW
Fox
I
I
Krizaj
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002540:Cerebral Cortex / Q000378:metabolism*; D002846:Chromatography, Affinity; D054501:Group II Phospholipases A2; D007457:Iodine Radioisotopes; D007700:Kinetics; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010741:Phospholipases A / Q000493:pharmacokinetics*; D054467:Phospholipases A2; D011869:Radioligand Assay; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D054507:Receptors, Phospholipase A2; D013552:Swine; D014757:Viper Venoms / Q000493:pharmacokinetics*
D002147:Calmodulin; D007457:Iodine Radioisotopes; C516360:PLA2R1 protein, human; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D054507:Receptors, Phospholipase A2; D014757:Viper Venoms; D010741:Phospholipases A; D054501:Group II Phospholipases A2; D054467:Phospholipases A2; C060161:ammodytoxin C