Preparation of biologically active analogs of serum low density lipoprotein.
A method for the preparation of stable and water-soluble analogs of low density lipoprotein (LDL) is presented. The experimental protocols start with the preparation of a cholesteryl ester/phospholipid microemulsion by a combined injection-sonication procedure and delipidation of apoprotein B (apoB) with sodium deoxycholate (NaDOC). The association of lipid microemulsion and NaDOC-solubilized apoB is achieved by incubation and sonication of the components above the melting point of the cholesteryl ester. The reconstituted model LDL (m-LDL) proved to be quite homogeneous both with respect to particle size and composition. Negative-stain electron microscopy shows spherical particles with a mean diameter of 21 nm. The mean density of the reconstituted LDL was 1.07 g/ml as determined by sucrose density gradient centrifugation. The reconstituted LDL retained its beta-mobility on agarose gel electrophoresis, and sodium dodecyl sulfate (SDS)-gel electrophoresis showed no degradation of apoB during the reconstitution procedures. Studies of biological activity showed that the m-LDL particles are bound, incorporated, and degraded by human fibroblasts in a way similar to native LDL. The reconstituted m-LDL has potential use for metabolic, physiochemical, and enzymatic studies of lipoproteins.
D001053:Apolipoproteins; D001055:Apolipoproteins B; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D002788:Cholesterol Esters / Q000032:analysis; D002850:Chromatography, Gel; D003840:Deoxycholic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D004655:Emulsions; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D008077:Lipoproteins, LDL / Q000032:analysis / Q000138:chemical synthesis* / Q000378:metabolism; D010713:Phosphatidylcholines / Q000032:analysis; D013010:Sonication
D001053:Apolipoproteins; D001055:Apolipoproteins B; D002788:Cholesterol Esters; D004655:Emulsions; D008077:Lipoproteins, LDL; D010713:Phosphatidylcholines; D003840:Deoxycholic Acid