Binding sites for calcium-activated neutral protease on erythrocyte membranes are not membrane phospholipids.
In order to explore the binding sites for calcium-activated neutral protease (CANP) with high calcium sensitivity (muCANP) on the inner surface of human erythrocyte membranes, we analyzed the binding of muCANP to two kinds of membranes modified by treatment with phospholipase C or Triton X-100. Binding analyses were performed using an immunoblot technique. The amount of muCANP bound to phospholipase C-treated inside-out vesicles was essentially the same as that bound to untreated inside-out vesicles. It was also observed that muCANP binds to Triton X-100-treated membranes, in which most of the integral proteins and glycerophospholipids are removed while the lining proteins remain intact. In both types of modified membrane, the bound muCANP was rapdily converted to an active form by autolysis at physiological free Ca2+ concentrations. These results indicate that the binding sites for muCANP on the inner surface of erythrocyte membranes consist of components other than membrane phospholipids. In addition, it is suggested that one of the binding sites for muCANP is some lining protein.
Department of Biochemistry, Tokyo Metropolitan Institute of Gerontology, Japan.
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Inomata
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Saito
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Kon
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Kawashima
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000097:blood / Q000494:pharmacology; D002154:Calpain / Q000097:blood* / Q000302:isolation & purification; D003902:Detergents / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000378:metabolism*; D006801:Humans; D008563:Membrane Lipids / Q000097:blood; D008565:Membrane Proteins / Q000097:blood*; D009132:Muscles / Q000201:enzymology; D017830:Octoxynol; D010743:Phospholipids / Q000097:blood; D011092:Polyethylene Glycols / Q000494:pharmacology; D011817:Rabbits; D010738:Type C Phospholipases / Q000494:pharmacology
D003902:Detergents; D008563:Membrane Lipids; D008565:Membrane Proteins; D010743:Phospholipids; D011092:Polyethylene Glycols; D017830:Octoxynol; D010738:Type C Phospholipases; D002154:Calpain; D002118:Calcium