In this report we describe a new method which is useful for measuring hydrophobic interactions between aliphatic hydrocarbon chains and proteins in aqueous environment. The method is based on partition of proteins in an aqueous two-phase system containing dextran and poly(ethy1ene glycol) and different fatty acid esters of poly(ethy1ene glycol). The partition is measured under conditions where contributions from electrostatic interactions are eliminated. The difference in partition of proteins in phase systems with and without hydrocarbon groups bound to poly(ethy1ene glycol), Alog K, where K is the partition coefficient, is taken as a measure of hydrophobic interaction. dlog K varies with size of hydrocarbon chain and type of protein. The length of the aliphatic chain should be greater than 8 carbon atoms in order to get a measurable effect in terms of dlog K. Bovine serum albumin, P-lactoglobulin, hemoglobin and myoglobin have been shown to have different affinities for palmitic acid ester of poly(ethy1ene glycol). No hydrophobic effect could be observed for ovalbumin, cytochrome c or a-chymotrypsinogen A.Hydrophobic interactions are known to be of essential importance for the stability, conformation and function of biological macromolecules and cells. It has therefore been the subject of several investigations both theoretical and experimental, and attempts have been made to establish a 'hydrophobic scale' within a given class of biological substances [l-41.Such a classification should be of help in understanding the mechanism of hydrophobic interactions as well as in preparative work. In the case of small molecules, hydrophobicity has been measured mainly in terms of their solubility in organic liquids [2,4], or their partition between an aqueous and an organic phase [5,6]. The results are then used to explain the interactions of such molecular groups within a protein or the affinity of hydrophobic ligands for proteins.We propose a new approach to measure hydrophobicity of macromolecules and cells as well as small molecules, based on partition in aqueous two-phase systems containing dextran and poly(ethy1ene glycol)With this method we can directly compare a variety of compounds in one and the same system, which further has the advantage that both phases are rich in water, 80-95% (wlw). These systems consists of a dextran-rich lower-phase and a poly(ethy1ene 171. glycol)-rich upper-phase. If a ligand is bound to one of these polymers it will be partitioned in the same way as the unsubstituted polymer. Thus by introducing a polymer-bound ligand in the system it is possible to change the relative affinity of a partitioned substance, e.g. a protein, for the phases. This has already been demonstrated [8,9]. Similarly when the phase system contains a polymer-bound hydrophobic ligand the corresponding phase will be more apolar.The partition coefficient of a protein is defined as K = c,/c,, where c, and c, are its concentrations at equilibrium in the upper and the lower phase respectively. Since, at equilibrium, the...
