Extraction of the apo B cluster from human low density lipoprotein with Tween 80.
The interaction of human serum low density lipoprotein with a non-ionic detergent, Tween 80, was investigated by sedimentation velocity measurements and Sepharose CL-4B gel chromatography. The properties of Tween 80-extracted apoprotein (apo B) were studied by sedimentation velocity and sedimentation equilibrium measurements, gel chromatography, electron microscopy and chemical cross-linking. Tween 80, at low concentrations, increased the sedimentation coefficient of LDL in 3% NaCl to a plateau value. At the same time, the effective Stokes radius of LDL expressed as the partition coefficient in gel chromatography became larger. The effect was reversed in part after protease treatment. A further increase of Tween 80 concentration caused the all-or-none dissociation of the entire mass of apo B from LDL. The remaining lipid core of LDL was not destroyed by detergent at 25 degrees C, but it was at 35 degrees C. The Tween 80-extracted apo B was purified by gel chromatography and its molecular weight and partial specific volume were determined by sedimentation equilibrium analysis in H2O and D2O buffers. It formed a complex of 1,200,000 molecular weight with Tween 80, with f/fo = 2.2. The molecular weight of the protein part of the complex was estimated to be about 500,000, corresponding to the total apo B in an LDL molecule. This result was confirmed by chemical cross-linking of apo B in Tween 80. Under an electron microscope the apo B-Tween 80 complex appeared as a flexible string, a little less than 1,000 A in length and 50 to 60 A in width.
D001053:Apolipoproteins / Q000097:blood*; D001055:Apolipoproteins B; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008077:Lipoproteins, LDL / Q000097:blood*; D008854:Microscopy, Electron; D008970:Molecular Weight; D011136:Polysorbates; D014461:Ultracentrifugation
D001053:Apolipoproteins; D001055:Apolipoproteins B; D008077:Lipoproteins, LDL; D011136:Polysorbates