Isolation and characterization of plasma membrane from Acanthamoeba culbertsoni.

ABSTRACT

A rapid method for preparation of plasma membrane from Acanthamoeba culbertsoni involving toluene treatment followed by lithium bromide extraction is described. In the plasma membrane preparation, 5'-nucleotidase, Na+ + K+ -ATPase, Mg2+ -ATPase and glucose-6-phosphatase activities were enriched. The membrane preparation was free from nucleic acid, cytochrome P-450 and cytochrome b5. Amino acid (14C-Ieucine) was not incorporated in the plasma membrane in 2 min. Succinic dehydrogenase was not detectable in the plasma membrane preparation. The molar ratio of cholesterol and phospholipids was 0·95 which is characteristics for plasma membranes. Under electronmicroscopy the preparation was homogenous without any other component of the cell. Plasma membrane proteins and glycoproteins were separated on acrylamide gel electrophoresis.