Purification and characterization of apolipoprotein J.
Apolipoprotein J (apoJ), a unique 70-kDa component of high density lipoproteins in human plasma, consists of two disulfide-linked subunits designated apoJ alpha (34-36 kDa), and apoJ beta (36-39 kDa) which share pI values of 4.9-5.4 and which are recognized by a monoclonal antibody (mAb) 11. ApoJ and its subunits were purified to homogeneity from plasma by a combination of immunoaffinity chromatography, using mAb11 linked to Affi-Gel, and reverse-phase high performance liquid chromatography. ApoJ alpha and apoJ beta are both glycoproteins. When deglycosylated, the molecular mass of apoJ alpha is 24 kDa and that of apoJ beta is 28 kDa, suggesting that approximately 30% of the mass of each subunit is carbohydrate. The amino acid compositions of apoJ alpha and apoJ beta are very similar; however, the sequences of the first 30-amino acid residues are distinct. A comparison of peptide maps suggests that apoJ alpha and apoJ beta are not identical but share limited regions of homology. This possibility is supported by immunochemical data. Five additional mAb specific for apoJ were characterized. One of the mAb, like mAb11, reacts with both apoJ alpha and apoJ beta; the others react with apoJ alpha only. All mAb, including those which recognize both apoJ alpha and apoJ beta and those which recognize apoJ alpha only, immunoprecipitate a approximately 50-kDa protein synthesized from a liver mRNA template translated in a rabbit reticulocyte lysate. We propose that the apoJ alpha and apoJ beta subunits, which have limited homology, are derived by proteolytic cleavage of a common precursor.
de Silva
H V
HV
Department of Pharmacology and Cell Biophysics, College of Medicine, University of Cincinnati, Ohio 45267-0575.
Stuart
W D
WD
Park
Y B
YB
Mao
S J
SJ
Gil
C M
CM
Wetterau
J R
JR
Busch
S J
SJ
Harmony
J A
JA
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001053:Apolipoproteins / Q000097:blood* / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D051152:Clusterin; D015180:Electrophoresis, Gel, Two-Dimensional; D006023:Glycoproteins; D006801:Humans; D008075:Lipoproteins, HDL / Q000097:blood* / Q000302:isolation & purification; D046911:Macromolecular Substances; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000302:isolation & purification
D000596:Amino Acids; D001053:Apolipoproteins; C495242:CLU protein, human; D051152:Clusterin; D006023:Glycoproteins; D008075:Lipoproteins, HDL; D046911:Macromolecular Substances; D018832:Molecular Chaperones; D010446:Peptide Fragments