Amphiphilic and hydrophilic molecular forms of acetylcholinesterase in membranes derived from sarcoplasmic reticulum of skeletal muscle.
Native molecular forms of acetylcholinesterase (AChE) present in a microsomal fraction enriched in SR of rabbit skeletal muscle were characterized by sedimentation analysis in sucrose gradients and by digestion with phospholipases and proteinases. The hydrophobic properties of AChE forms were studied by phase-partition of Triton X-114 and Triton X-100-solubilized enzyme and by comparing their migration in sucrose gradient containing either Triton X-100 or Brij 96. We found that in the microsomal preparation two hydrophilic 13.5 S and 10.5 S forms and an amphiphilic 4.5 S form exist. The 13.5 S is an asymmetric molecule which by incubation with collagenase and trypsin is converted into a 'lytic' 10.5 S form. The hydrophobic 4.5 S form is the predominant one in extracts prepared with Triton X-100. Proteolytic digestion of the membranes with trypsin brought into solution a significant portion of the total activity. Incubation of the membranes with phospholipase C failed to solubilize the enzyme. The sedimentation coefficient of the amphiphilic 4.5 S form remained unchanged after partial reduction, thus confirming its monomeric structure. Conversion of the monomeric amphiphilic form into a monomeric hydrophilic molecule was performed by incubating the 4.5 S AChE with trypsin. This conversion was not produced by phospholipase treatment.
Cánovas-Muñoz
M D
MD
Departamento de Bioquímica y Biología Molecular, Facultad de Biología, Universidad de Murcia, Spain.
Campoy
F J
FJ
Muñoz-Delgado
E
E
Vidal
C J
CJ
D000110:Acetylcholinesterase / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D003902:Detergents / Q000494:pharmacology; D007461:Iodoacetates / Q000494:pharmacology; D019807:Iodoacetic Acid; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology*; D009132:Muscles / Q000201:enzymology*; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D012995:Solubility
D003902:Detergents; D007461:Iodoacetates; D007527:Isoenzymes; D011092:Polyethylene Glycols; D017830:Octoxynol; C010615:Nonidet P-40; D000110:Acetylcholinesterase; D019807:Iodoacetic Acid