Identification of proteins associated with apolipoprotein A-I-containing lipoproteins purified by selected-affinity immunosorption.
The isolation of apolipoprotein A-I-containing lipoproteins [Lp(A-I)] by selected-affinity immunosorption minimizes the loss of associated proteins that occurs during the isolation of high-density lipoproteins (HDL) by sequential ultracentrifugation. We have used two-dimensional gel electrophoretic analysis to separate the proteins associated with Lp(A-I). Using a combination of amino acid sequencing of transblotted proteins and Western blotting with specific antisera, we have identified a number of associated proteins. The positions of the apolipoproteins (apo) A-I, A-II, A-IV, C-III, D, and E were located on the gels. Lecithin-cholesterol acyltransferase and cholesteryl ester transfer protein were identified in association with Lp(A-I) to a greater extent than found associated with HDL after centrifugation. In addition to those proteins previously identified in association with HDL, we detected a number of plasma proteins associated with Lp(A-I), namely, fibrinogen, haptoglobin, proline-rich protein (C4b-binding protein), and apolipoprotein J (SP40,40 sulfated glycoprotein). The co-isolation of these proteins with Lp(A-I) does not appear to be an artifact in that they have very low affinity for a sham column containing covalently bound preimmune goat IgG in place of the anti-apoA-I IgG. These findings suggest that in addition to apolipoproteins that exist largely in association with lipoproteins there is another class of proteins which exist in lipoprotein-associated form and in the dispersed state. Detection and identification of these lipoprotein-associated proteins may aid in the mechanistic determination of a number of observed functions attributed to HDL.
Kunitake
S T
ST
Cardiovascular Research Institute, University of California, San Francisco 94143.
Carilli
C T
CT
Lau
K
K
Protter
A A
AA
Naya-Vigne
J
J
Kane
J P
JP
D000595:Amino Acid Sequence; D016632:Apolipoprotein A-I / Q000032:analysis*; D001798:Blood Proteins / Q000032:analysis*; D015153:Blotting, Western; D002846:Chromatography, Affinity; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D006801:Humans; D008074:Lipoproteins / Q000737:chemistry* / Q000302:isolation & purification; D008297:Male; D008969:Molecular Sequence Data
D016632:Apolipoprotein A-I; D001798:Blood Proteins; D008074:Lipoproteins