Demonstration of a high density lipoprotein (HDL)‐binding protein in Hep G2 cells using colloidal gold‐HDL conjugates

Abstract

Solubilized membrane proteins of Hep G2 cells were electrophoretically separated on polyacrylamide gels and electrotransferred onto nitrocellulose paper. Overlaying the nitrocellulose with human high density lipoproteins conjugated to colloidal gold revealed the presence of a single protein band with an apparent molecular mass of 80 kDa. Binding of the conjugates to this protein was specific for high density lipoproteins in as much as it was effectively displaced by an excess of unlabelled high density lipoproteins but not by a similar excess of unlabelled low density lipoproteins. Binding was not dependent on Ca*+ as 10 mM EDTA had no effect. The binding activity of the solubilized membranes was increased by incubating the cells with non-lipoprotein cholesterol. This was detected on electroblots and quantified with a new dot blot assay using the colloidal gold-high density lipoprotein conjugates.

Retroendocytosis of high density lipoproteins by the human hepatoma cell line, HepG2.

“…2032 Recently, we have reported the presence of an 80 kDa protein in HepG2 cells, which specifically bound colloidal gold-HDL 3 and was up-regulated by cholesterol. 12 Such findings are in contrast to the welldocumented down-regulation of the LDL receptor by cholesterol. 1441 In the present study, the cholesterolinduced increase in the specific binding of 125 I-HDI_3 at 4°C was reflected in a proportional increase in both the amount of label internalized and that released during the chase.…”

“…H igh density lipoproteins (HDL) stimulate the removal of cholesterol from several types of peripheral cells In culture 12 and are thought to play a similar role in vivo. 34 They have, therefore, been implicated in the transport of cholesterol from peripheral tissues to the liver for disposal or re-utilization, a process called reverse cholesterol transport.…”

Studies on the proteins involved in the interaction of high‐density lipoprotein with isolated human small intestine epithelial cells

“…Several attempts have been undertaken to identify determinants in both HDL [5] and its putative receptor [7][8][9][10]. These studies have, however, not resulted in a consensus for the identities of specific determinants involved in this interaction.…”

“…Proteins which specifically bind ape A-I and HDL have been isolated and partially characterized from a number of cells, These proteins may represent the putative HDL receptor or parts thereof [7][8][9][10]. One of these proteins, isolated from human placenta by affinity chromatography, specifically bound to ape A-I and was designated as ape A-I-binding protein (ape A-I-BP) [9].…”

High density lipoprotein-binding proteins in porcine liver. Isolation and histological localization.

“…In rat liver, HDL-binding proteins of 78 kd 14 and 100 and 120 kd 19 have been described, while in the rat hepatoma cell line Fao, a 110-kd HDL-binding protein has been detected. 15 In the human hepatoma cell line HepG2, both an 80-kd 32 and a 110-kd 17 HDL-binding protein have been reported. There are no reports on the functional differences between these proteins.…”