Effects of aliphatic fatty acids on the binding of Phenol Red to human serum albumin

Abstract

Binding of Phenol Red to human serum albumin at pH 7.0 was studied by ultrafiltration (n1 = 1, K1 = 3.9 X 1-(4) M-1, n2 = 5, K2 = 9.6 X 10(2) M-1). The presence of 1 mol of octanoate or decanoate per mol of albumin caused a decrease in dye binding (dye/protein molar ratio 1:1), which, in contrast with additional fatty acid, was very pronounced: 1-8 mol of palmitate or stearate resulted in a small, and apparently linear, displacement of Phenol Red. The displacement effect of 1-5 mol of oleate, linoleate or linolenate per mol of albumin was comparable with that of the equimolar concentrations of palmitate or stearate. A higher molar ratios the unsaturated acids caused a drastic decrease in dye binding. The different Phenol Red-displacement effects of low molar ratios of medium-chain and long-chain fatty acids indicate that these acids have different high-affinity binding sites. In accordance with this proposal, low concentrations of stearate had only a small effect on the Phenol Red-displacement effect of octanoate. Phenol Red-binding curves in the presence of 1 mol of octanoate, 8 mol of stearate and 6 or 7 mol of linolenate per mol of albumin respectively indicated that the dye and the fatty acids do not complete for a common primary binding site. In contrast, a secondary Phenol Red-binding site could be identical with the primary octanoate-binding site. Furthermore, the primary Phenol Red-binding site could be the same as a secondary linolenate-binding site. Assignment of the different primary binding sites for Phenol Red and for medium-chain and long-chain fatty acids to a model of the secondary structure of albumin is attempted.

Relations between high-affinity binding sites of markers for binding regions on human serum albumin

“…Therefore, as a part of the present study, the individual binding of warfarin, digitoxin, diazepam, salicylate and Phenol Red to defatted human serum albumin was studied. The binding parameters n and K were calculated by using the double-reciprocal-plot method of Klotz (1946) as previously described (Kragh-Hansen, 1981a (1974) found n, = 0.95 and K1 = 2.31 x 105M-1 (equilibrium-dialysis and ultrafiltration studies with media containing 0.2M-phosphate buffer, pH 7.4, at 37°C). Sudlow et al (1975) reported n1 = 0.9 and K1 = 2.5 x 105 M-1 (fluorimetry, 0.1 M-sodium phosphate/0.9% NaCl, pH7.4, at 22°C).…”

Application of phenol red as a marker ligand for bilirubin binding site at subdomain IIA on human serum albumin

“…Thus, the decrease of binding of PhRed in the PHSerum could be only partially due to displacement by bilirubin. According to the literature the binding of PhRed may be reduced by the presence of different aliphatic fatty acids [22] or may be displaced by Cl - [20], which also bind to HSA.…”

“…The interaction of serum albumin with various dyes such as PhRed [16][17][18][19][20][21][22][23][24][25][26][27][28][29], bromophenol blue [29][30][31][32], bromocresol green [29,33], bromocresol purple [34], indocyanine green [29], sulfobromophtalein [29] and rose bengal [29,35] was studied by numerous investigators. The results obtained by Kamisaka et al [29] from the competitive binding of bilirubin and some dyes showed that the dyes compete for the common binding sites occupied by bilirubin, but the binding specificity and mode of interaction depend on the structure of the dyes and the site where the displacement occurs.…”

Binding of daunorubicin to human serum albumin using molecular modeling and its analytical application

“…HSA can bind and carry through the bloodstream many drugs, including anticoagulants, tranquilizers, and general anesthetics [5,6] are transported in the blood while bound to albumin, which are poorly soluble in water. It has been shown that the distribution, free concentration and the metabolism of various drugs can be significantly altered as a result of their binding to HSA [7]. Drug interactions at protein binding level will in most cases significantly affect the apparent distribution volume of the drugs and also affect the elimination rate of drugs.…”