Studies on human serum high-density lipoproteins (HDL). IV. Isolation of lipoprotein families after incubation of HDL.
The high-density lipoproteins (HDL) of human serum appear to be unstable and easily exposed to chemical changes during isolation. In earlier studies we have isolated and purified HDL subfractions either in the presence of an SH-blocking agent, DTNB, or in the cold. By both procedures reproducible lipoprotein subfractions could be recovered by hydroxyl apatite column chromatography at the elution steps 0.03-0.05 mol/l (subfraction II) and 0.05-0.15 mol/l phosphate buffer (subfraction III). The protein moiety of both lipoprotein subfractions contained polypeptides A-I , A-II, thin line (TL), C-I and C-II, and the protein moiety of subfraction III contained also C-III. The incubation at 37 degrees C of these HDL subfractions gave reproducible daughter lipoprotein fractions that could be recovered by subsequent rechromatography on hydroxyl apatite. At each of the elution steps 0.05-0.075 mol/l and 0.075-0. mol/l one daughter fraction was recovered, the protein moiety of which was composed of polypeptide A-I, as judged by polyacrylamide gel electrophoresis, immunodiffusion, and amino acid analysis. The incubation of parent subfractions II and III caused also the appearance at elution step 0.001-0.01 mol/l of a daughter lipoprotein fraction - lipoprotein A (Lp-A) - that was characterized by a protein moiety with polypeptides A-I and A-II in equal amounts. The 'release' of lipoprotein A-I (Lp-A-I) and Lp-A was shown to be due rather to the incubation than to the column chromatography as such. The chemical changes occurring during the incubation of HDL suggested a degradation of phosphatidylcholine (PC) to lysophosphatidylcholine (lyso-PC) and glycerylphosphorylcholine (GPC). It is suggested that the degradation of PC might interfere with the interaction between the lipoprotein families composing HDL.
D000596:Amino Acids / Q000097:blood; D002784:Cholesterol / Q000302:isolation & purification; D002850:Chromatography, Gel; D003208:Concanavalin A; D004591:Electrophoresis, Polyacrylamide Gel; D005997:Glycerylphosphorylcholine / Q000302:isolation & purification; D006801:Humans; D006882:Hydroxyapatites; D008055:Lipids / Q000097:blood; D008075:Lipoproteins, HDL / Q000097:blood* / Q000302:isolation & purification; D008297:Male; D010713:Phosphatidylcholines / Q000302:isolation & purification; D010714:Phosphatidylethanolamines / Q000302:isolation & purification; D012685:Sepharose; D013109:Sphingomyelins / Q000302:isolation & purification
D000596:Amino Acids; D006882:Hydroxyapatites; D008055:Lipids; D008075:Lipoproteins, HDL; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D013109:Sphingomyelins; D003208:Concanavalin A; D005997:Glycerylphosphorylcholine; D012685:Sepharose; D002784:Cholesterol