Protein-lipid interactions in antipodal plasma membranes of rat colonocytes.
The isolation of apical membranes from rat proximal colonic epithelial cells is described. Differential centrifugation yielded a 'crude' membrane fraction which was further purified using sucrose density centrifugation. The final membrane fraction was enriched 20-28-fold over homogenate in alkaline phosphatase and cysteine-sensitive alkaline phosphatase specific activities. Lipid-protein interactions and lipid dynamics examined in apical and basolateral membranes prepared from colonocytes demonstrated: (1) apical membrane, as assessed by steady-state fluorescence polarization studies have a low lipid fluidity; (2) colonic basolateral membranes possess a greater lipid fluidity than apical membranes; (3) compositional differences in these antipodal membranes appear to explain these differences in lipid fluidity; (4) fluorescence polarization studies using diphenylhexatriene detect a thermotropic transition at 21-23 degrees C in apical membranes and liposomes prepared from lipid extracts of these membranes; (5) alkaline phosphatase and L-cysteine-sensitive alkaline phosphatase activities appear to be functionally dependent on the physical state of the apical membrane's lipid.
D000251:Adenosine Triphosphatases / Q000032:analysis; D000818:Animals; D002458:Cell Fractionation; D002462:Cell Membrane / Q000201:enzymology / Q000648:ultrastructure; D003106:Colon / Q000201:enzymology / Q000648:ultrastructure*; D008297:Male; D008563:Membrane Lipids / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D010088:Oxidoreductases / Q000032:analysis; D010744:Phosphoric Monoester Hydrolases / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains; D013816:Thermodynamics
D008563:Membrane Lipids; D008565:Membrane Proteins; D010088:Oxidoreductases; D010744:Phosphoric Monoester Hydrolases; D000251:Adenosine Triphosphatases