Characterization of a membrane-associated receptor from rat sinusoidal liver cells that binds formaldehyde-treated serum albumin.

S Horiuchi,K Takata,Y Morino

doi:10.1016/s0021-9258(18)89757-5

Abstract

When treated with formaldehyde, serum albumin is known to be taken up and degraded by sinusoidal liver cells via adsorptive endocytosis. The present study aimed at characterization and identification of the membrane-associated receptor on rat sinusoidal liver cells. Kinetic studies of binding of 125I-labeled formaldehyde-treated serum albumin (125I-f-Alb) with the membranes of sinusoidal liver cells demonstrated the presence of specific, high-affinity, saturable membrane-bound receptors with an apparent Kd = 8 micrograms of f-Alb/ml and the optimal pH at around 8.0. The 125I-f-Alb binding to the membranes was not inhibited by either native albumin, asialofetuin, methylamine-treated alpha 2-macroglobulin, mannan, or immune complexes. The binding process exhibited independence of calcium and susceptibility both to heat treatment and to destruction by proteases. The binding was inhibited by concanavalin A and the inhibition was effectively reversed by the presence of alpha-methyl-D-glucoside, a haptenic inhibitor for this lectin, indicating the glycoprotein nature of the receptor. The binding protein was extracted from the membrane preparations with octyl beta-D-glucopyranoside and immunoprecipitated by anti-ligand antibody as a complex with the ligand. Sodium dodecyl sulfate-gel electrophoresis of the immunoprecipitate revealed two polypeptide chains with molecular weights of approximately 53,000 and 30,000, respectively.

Highlights

Since it isgenerally accepted that the specificity of receptormediated endocytosis residesexclusively on the initial binding step between a ligand and its receptor [12], we directed our attentiontotheinitialbinding process by analyzing the interaction of f-Albwith themembranefractions isolated from sinusoidal liver cells
Binding of lZ5I-f-Albto Sinusoidal Liver Cell MembranesFig. 1 showsthetimecourse for binding of 'T-f-Albto membranes isolated from rat sinusoidal iver cells
Albumin preparations which were S-carboxymethylated or S-carboxamidomethylated in the presenocfe6 M guanidine hydrochloride were ineffective at a concentration of 200 pg/ml. These findings indicate that the drastic denaturation of albumin molecules is not requiredfor the specific recognition by the membrane-associatedreceptor

Summary

The present paper describes first some characteristics of

As early as 1965, Mego and McQueen [1] found that for- binding of Iz5I-labeled f-Alb to sinusoidal cell membranes, maldehyde-treated 1311-serumalbumin was rapidly cleared and, second, the identification of the binding protein in the from theblood stream and taken buypthe liver wheninjected nonionic detergent extract of sinusoidal cell membranes as a intravenously into mice [1].Subsequent studies by Nilsson binding protein-ligandcomplex with monospecific anti-ligand and Berg( 2 ) showed that f-Alb’ was exclusively taken up and antibody. degraded by sinusoidal liver cells both in vivo and in uitro.

Thekineticstudies on in uitro uptake using isolatedrat

RESULTS

Experiment A None

DISCUSSION

Slice Number