Physicochemical behaviour and structural characteristics of membrane-bound acetylcholinesterase from Torpedo electric organ. Effect of phosphatidylinositol-specific phospholipase C.
Quantitative solubilization of the phospholipid-associated form of acetylcholinesterase (AChE) from Torpedo electric organ can be achieved in the absence of detergent by treatment with phosphatidylinositol-specific phospholipase C (PIPLC) from Staphylococcus aureus [Futerman, Low & Silman (1983) Neurosci. Lett. 40, 85-89]. The sedimentation coefficient on sucrose gradients of AChE solubilized in detergents (DSAChE) varies with the detergent employed. However, the coefficient of AChE directly solubilized by PIPLC is not changed by detergents. Furthermore, PIPLC can abolish the detergent-sensitivity of the sedimentation coefficient of DSAChE purified by affinity chromatography, suggesting that one or more molecules of phosphatidylinositol (PI) are co-solubilized with DSAChE and remain attached throughout purification. DSAChE binds to phospholipid liposomes, whereas PIPLC-solubilized AChE and DSAChE treated with PIPLC do not bind even to liposomes containing PI. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis shows that PIPLC-solubilized AChE, like unmodified DSAChE, is a catalytic subunit dimer; electrophoresis in the presence of reducing agent reveals no detectable difference in the Mr of the catalytic subunit of unmodified DSAChE, of AChE solubilized by PIPLC and of AChE solubilized by Proteinase K. The results presented suggest that DSAChE is anchored to the plasma membrane by one or more PI molecules which are tightly attached to a short amino acid sequence at one end of the catalytic subunit polypeptide.
Futerman
A H
AH
Fiorini
R M
RM
Roth
E
E
Low
M G
MG
Silman
I
I
D000110:Acetylcholinesterase / Q000378:metabolism*; D000818:Animals; D002462:Cell Membrane / Q000201:enzymology; D002499:Centrifugation, Density Gradient; D003902:Detergents / Q000494:pharmacology; D004557:Electric Organ / Q000187:drug effects / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000494:pharmacology; D008081:Liposomes / Q000378:metabolism; D008954:Models, Biological; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D010740:Phospholipases / Q000494:pharmacology; D010727:Phosphoric Diester Hydrolases / Q000494:pharmacology*; D014101:Torpedo / Q000378:metabolism*
D003902:Detergents; D008081:Liposomes; D010740:Phospholipases; D000110:Acetylcholinesterase; D010727:Phosphoric Diester Hydrolases; D054801:Phosphoinositide Phospholipase C; D010450:Endopeptidases; D043265:Phosphatidylinositol Diacylglycerol-Lyase