Functional characterization of albumin binding to the apical membrane of OK cells

Abstract

We characterized binding of albumin to the apical membrane of opossum kidney (OK) cells using fluorescein isothiocyanate (FITC)-albumin (i.e., bovine serum albumin, BSA) as substrate. Functional analysis of binding data showed one specific binding site characterized by half-maximal binding (Michaelis constant, (Km) at 20 mg/l (300 nmol/l) and maximal binding capacity (Bmax) of 0.61 microgram/mg cellular protein. Excess of unlabeled albumin (BSA) inhibited binding at low concentrations of FITC-albumin completely but only partially at high concentrations. FITC-albumin binding was reversible and pH dependent. Km increased about sixfold when pH decreased from 7.4 to 5.0. The inhibitory effects of conalbumin, alpha-lactalbumin, and transferrin were significantly smaller compared with BSA. We conclude that OK cells express a high-affinity binding site for albumin on the apical membrane. This binding site is pH sensitive, binds albumin in the physiological range, and could be responsible for the effective receptor-mediated reabsorption of albumin in the proximal tubule.