The binding of indole compounds to bovine plasma albumin. Effects of potassium chloride, urea, dioxane, and glycine.

By the use of a rapid dialysis equilibrium technique, the binding of skatole and acetyl-L-tryptophan with bovine albumin has been studied at various concentrations of KCl, dioxane, urea, and glycine, at different pH values, and at different temperatures. Binding was found to be principally at one site with an association constant much higher than at any secondary sites. The primary association constant, A’, for skatole at pH 6.3, 0.1 M KCl, was 4.2 x lo5 M-’ (4’) and 1.9 X lo5 M-’ (18’); for acetyl-L-tryptophan at the same conditions, k’1 was 3.5 X lo5 M-’ (4O) and 1.8 X lo5 M-’ (18’). A decrease in KC1 concentration from 0.1 to 0 decreased k’, for skatole to a value & and r’r; that in 0.1 M KCI. A decrease in KC1 concentration from 0.1 to 0.002 M, on the other hand, increased k’l for acetyl-L-tryptophan approximately 3-fold. Chloride was a competitive inhibitor of acetyl-Ltryptophan. The addition of either dioxane or urea reduced k’l for skatole and acetyl-L-tryptophan, whereas the addition of glycine had no effect on k’l. When added to urea solutions, glycine increased k’l. The activity coefficient of skatole, y, in urea, dioxane, or glycine-water mixtures and urea-glycine-water mixtures was determined by solubility measurements. The changes in k’, for skatole in dioxane at all concentrations studied and in urea up to 2 M could be approximately reconciled by the assignment 1 l-y2 for the ratio of the activity coefficients of the protein-skatole complex and the unassociated protein. The similarities in the effects of dioxane and urea (up to 2 M) on the association of the ligand have been taken to indicate that the primary influence of these solutes is on the apolar surfaces of the associating molecules. At approximately 4 M urea, k’l became larger at 18’ than at 4’, resulting in a crossing over of the temperature curves of k’l plotted as a function of urea concentration. Thermodynamic changes for the association were also estimated. Under the same conditions, AS’ was 5 to 15 calories per mol degree more negative for skatole than for acetyl-L-tryptophan. AS’ increased markedly at urea concentrations above 2 M urea and also increased, but at a