Preparation and properties of soluble, immunoreactive apoLDL.
Immunoreactive apo-low density lipoprotein (LDL), soluble in mildly alkaline buffers of low ionic strength, was prepared by attaching LDL to a DEAE-Sepharose column and eluting the lipids with a 0--2% (w/v) gradient of nonionic detergents. Brij-36T, Nonidet P-40, and Triton X-100 gave similar results. After washing the detergent from the column, the bound apoLDL was eluted with 1 M NaCl, pH 7.4, with recoveries up to 85%. This apoLDL could be dialyzed extensively against low ionic strength solutions, and remained soluble as long as the pH was above 7. Spectrophotometric analysis showed that less than 0.1% %w/v) of cholesterol or phospholipids and less than 1% (w/v) of detergent remained associated with the protein. The apoLDL cross-reacted with LDL against antisera prepared vs. intact LDL. Pore-gradient polyacrylamide gel electrophoresis, with SDS and urea, showed that this preparation was less aggregated than organic solvent extracted apolLDL and appeared to be made of oligomers of two monomeric subunits, one with molecular weight around 22,700 and a smaller one of approximately 8000. Isoelectric focusing showed that there also was charge heterogeneity in the soluble apoLDL.
D001053:Apolipoproteins / Q000276:immunology; D055598:Chemical Phenomena; D002621:Chemistry; D002850:Chromatography, Gel / Q000379:methods; D003902:Detergents; D004586:Electrophoresis / Q000379:methods; D006801:Humans; D005779:Immunodiffusion; D007525:Isoelectric Focusing; D008077:Lipoproteins, LDL / Q000097:blood* / Q000276:immunology; D046911:Macromolecular Substances; D012995:Solubility
D001053:Apolipoproteins; D003902:Detergents; D008077:Lipoproteins, LDL; D046911:Macromolecular Substances