Roles of lipids and proteins in the Ca2+-PO4-induced aggregation of cytoskeleton-free erythrocyte vesicle membranes.
The roles of lipids and proteins in Ca2+-PO4-induced membrane aggregation were investigated. Cytoskeleton-free vesicles derived from intact human and rabbit erythrocytes (HEves and REves, respectively) were employed as a model system. The HEves and REves have a simplified membrane protein composition [band 3 proteins and glycoproteins PAS-1, -2, and -3 (HEves)] and normal lipid composition. Optimal experimental conditions for pH, [PO4], and [CaCl2] were determined for quantitatively examining the dynamics and extent of HEves and REves aggregation, measured turbidimetrically. The aggregation process was found to be quite sensitive to small changes in pH and [PO4] and much less sensitive to [CaCl2]. The roles of membrane proteins in vesicle aggregation were examined by selectively modifying the proteins enzymatically. The roles of lipids were studied by using sonicated lipid vesicles [small unilamellar vesicles (SUVs)] made from Dodge ghost lipid extracts. Enzymatic treatment with trypsin, chymotrypsin, or Pronase had no effect on either the rates or the extent of vesicle aggregation (2-min incubation period). Neuraminidase treatment reduced both factors by approximately 20%. SUVs aggregated with Ca2+-PO4 in a way which depended on the PO4/lipid ratio. Together the results suggest the following: (1) PO4 is associated with the vesicle surface, involving the membrane lipids; (2) the vesicle + PO4 incubation time component of the PO4 effect is eliminated by enzymatically modifying the vesicle membrane proteins; (3) qualitative, rather than quantitative, properties of sialic acid containing molecules affect vesicle aggregation; and (4) with the exception of the incubation time effect, membrane proteins seem neither to promote nor to inhibit Ca2+-PO4-induced HEves or REves aggregation.
D002122:Calcium Chloride / Q000494:pharmacology*; D004910:Erythrocyte Membrane / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D008563:Membrane Lipids / Q000097:blood*; D008565:Membrane Proteins / Q000097:blood*; D009391:Nephelometry and Turbidimetry / Q000379:methods; D010710:Phosphates / Q000494:pharmacology*
D008563:Membrane Lipids; D008565:Membrane Proteins; D010710:Phosphates; D002122:Calcium Chloride