Immunologic comparison of the conformations of apolipoprotein B. Investigation of methodologies for the reconstitution of delipidated and denatured apolipoprotein B with nonionic surfactants.
Immunologic probes have been used to examine the conformation of apolipoprotein B (apo-B) as it exists within native low density lipoprotein (LDL) after lipid displacement with Triton X-100 and after denaturation with guanidine hydrochloride organic solvent delipidation and reconstitution with Triton X-100. Antigenic expression was assayed in two systems: by using either Triton X-100 or bovine serum albumin to maintain protein solubility. Apo-B delipidated by lipid displacement using Triton X-100 was virtually identical to LDL-apo-B in both systems, as assayed by polyclonal antisera prepared in rabbits against either antigen. Thus the native antigenic sites are preserved, although the displacement of the lipid core of LDL drastically alters the physical properties of the particle. Apo-B delipidated by solvent extraction in guanidine was reconstituted with Triton X-100 by several methods, and the products were examined immunologically. One method yielded a product that resembled apo-B as delipidated with Triton X-100, although full reconstitution could not be achieved. Nevertheless, Triton promoted refolding of apo-B to reform partial native structure as judged immunologically. By using both physical and immunologic methods for assessing structure, it is clearly evident that the perceptions of the conformational states of reconstituted apo-B can be very different, and multiple criteria need to be used to assess lipoprotein reconstitution.
D000936:Antigen-Antibody Complex; D001055:Apolipoproteins B / Q000097:blood* / Q000276:immunology; D003902:Detergents; D006801:Humans; D007106:Immune Sera; D005779:Immunodiffusion; D007700:Kinetics; D017830:Octoxynol; D011092:Polyethylene Glycols; D011487:Protein Conformation; D011863:Radioimmunoassay
D000936:Antigen-Antibody Complex; D001055:Apolipoproteins B; D003902:Detergents; D007106:Immune Sera; D011092:Polyethylene Glycols; D017830:Octoxynol