title: A novel method for converting apolipoprotein B, the major protein moiety of human plasma low density lipoproteins, into a water-soluble protein.
abstract: We have developed a new technique to solubilize apolipoprotein B (ApoB) in aqueous solutions. The procedure involves stirring ApoB in 6 M guanidine/20 mM NH4Cl/NH4OH in the presence of cupric ammonia complexes at pH 9.7 for 20 h in N2, and then removing these reagents by a series of dialysis in N2. The resulting Cu(NH3)4(2)+-treated (Cu2+-treated) ApoB is soluble in aqueous buffers of pH above 8.3 or below 3. Parallel experiments carried out on control proteins, human albumin, alpha-lactalbumin, and insulin, indicated no change in molecular weight and no creation of a new NH2-terminal amino acid after Cu2+-treatment. By Edman degradation, the Cu2+-treated ApoB showed no detectable NH2-terminal amino acid. These results showed that the mechanism of Cu2+-solubilization of ApoB was not due to the cleavage of peptide bonds. Electrophoresis on urea-polyacrylamide gel, Cu2+-treated ApoB showed the same number of bands as the non-treated ApoB in the separating gels (7%) near the cathode, suggesting the heterogeneity of ApoB. In SDS-polyacrylamide gel (10%), the reduced and Cu2+-treated ApoB migrated with the similar mobilities to the monomer or dimer of human albumin. Antibodies raised against Cu2+-treated ApoB gave at least two immunoprecipitin lines against the Cu2+-treated ApoB as well as the non-treated guanidine-HCl-soluble ApoB, suggesting the presence of non-identical subunits.
mesh_terms: D000596:Amino Acids / Q000032:analysis; D001053:Apolipoproteins; D055598:Chemical Phenomena; D002621:Chemistry; D003300:Copper / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006146:Guanidines; D006801:Humans; D005779:Immunodiffusion; D008077:Lipoproteins, LDL / Q000097:blood; D008722:Methods; D012995:Solubility; D014867:Water
chemical_list: D000596:Amino Acids; D001053:Apolipoproteins; D006146:Guanidines; D008077:Lipoproteins, LDL; D014867:Water; D003300:Copper