Role of apolipoprotein D in the transport of bilirubin in plasma.
Apolipoprotein D (apo D) is a 30-kDa glycoprotein of unknown function that is associated with high-density lipoproteins (HDL). Because unconjugated bilirubin has been shown to bind apo D with a 0. 8:1 stoichiometry, we examined the contribution of this protein to transport of bilirubin in human plasma. Density gradient centrifugation analysis using physiological concentrations of [(14)C]bilirubin reveals that 9% of unconjugated bilirubin is associated with HDL, with the remaining pigment bound primarily to serum proteins (i.e., albumin). The percentage of total plasma bilirubin bound to HDL was found to increase proportionally with bilirubin concentration. Affinity of human apo D for bilirubin was determined by steady-state fluorescence quenching, with Scatchard analysis demonstrating a single binding site for unconjugated bilirubin with an affinity constant (K(a)) of approximately 3 x 10(7) M(-1). Incorporation of apo D into phosphatidylcholine vesicles had no effect on K(a), suggesting that a lipid environment does not alter the affinity of the protein for bilirubin. Using stopped-flow techniques, the first-order rate constant for bilirubin dissociation from apo D was measured at 5.4 s(-1) (half-time = 129 ms). Our findings indicate that HDL is the principal nonalbumin carrier of bilirubin in human plasma and further support the proposition that the affinity of HDL for bilirubin is primarily the result of binding to apo D.
Goessling
W
W
Department of Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA. wgoessling@bics.bwh.harvard.edu
Zucker
S D
SD
D000818:Animals; D001053:Apolipoproteins / Q000097:blood*; D053399:Apolipoproteins D; D001663:Bilirubin / Q000097:blood*; D002250:Carbon Radioisotopes; D002784:Cholesterol / Q000097:blood; D019826:Cholic Acid / Q000097:blood; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008075:Lipoproteins, HDL / Q000097:blood; D008297:Male; D010949:Plasma / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012709:Serum Albumin / Q000378:metabolism
D001053:Apolipoproteins; D053399:Apolipoproteins D; D002250:Carbon Radioisotopes; D008075:Lipoproteins, HDL; D012709:Serum Albumin; D002784:Cholesterol; D019826:Cholic Acid; D001663:Bilirubin