[Separation and purification of human apolipoproteins A-I and C-III by chromatofocusing].
Human very low density lipoprotein (VLDL) and high density lipoprotein (HDL) were isolated and purified by a process of combined dextran sulfate precipitation and density gradient ultracentrifugation. Chromatofocusing, which separates protein based on differences in isoelectric point, was used to separate apolipoprotein A-I (apoA-I) and apolipoprotein C-III from human HDL and VLDL, respectively. Discontinuous SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and analytical isoelectric focusing (IEF) were used to study the purity of different fractions. Both purified apoA-I and apoC-III showed single bands on SDS-PAGE at molecular weights of 28183 and 9400 Daltons, respectively. As determined by IEF in the presence of 8 mol/L urea, apoA-I had eight isoforms with pI of 5.66-5.87. The pI's of the three isoproteins of apoC-III (C-III0, C-III1 and C-III2) were 5.06, 4.88 and 4.72, respectively. Chromatofocusing, a new simple technique combining the high resolving power of IEF with the high capacity of ion-exchange column chromatography, is extremely valuable for large-scale purification of the major apolipoproteins of VLDL and HDL.
Cheng
B
B
Institute of Basic Medical Sciences, CAMS and PUMC, Beijing.
D016632:Apolipoprotein A-I / Q000302:isolation & purification*; D053305:Apolipoprotein C-III; D001056:Apolipoproteins C / Q000302:isolation & purification*; D002852:Chromatography, Ion Exchange / Q000379:methods; D006801:Humans; D007525:Isoelectric Focusing / Q000379:methods; D008075:Lipoproteins, HDL / Q000737:chemistry; D008079:Lipoproteins, VLDL / Q000737:chemistry
D016632:Apolipoprotein A-I; D053305:Apolipoprotein C-III; D001056:Apolipoproteins C; D008075:Lipoproteins, HDL; D008079:Lipoproteins, VLDL