Partitioning of proteins and thylakoid membrane vesicles in aqueous two-phase systems with hydrophobically modified dextran

Abstract

Dextrans were modified with hydrophobic groups, i.e. benzoyl and valeryl groups. Benzoyl dextran and valeryl dextran form aqueous two-phase systems with poly(ethylene glycol) (PEG) as well as with dextran. Two-phase systems are also formed between two valeryl dextrans with different degrees of substitution as well as between benzoyl dextran and valeryl dextran. Phase diagrams for aqueous two-phase systems composed of PEG-valeryl dextran, dextran-valeryl dextran, valeryl dextran-valeryl dextran and valeryl dextran-benzoyl dextran have been determined. The effects of these hydrophobic groups on the partitioning of amino acids, proteins and membrane vesicles in aqueous two-phase systems have been studied. In a PEG 8000-dextran T500 aqueous two-phase system containing only phosphate buffer β-galactosidase was partitioned mostly to the top phase. However, by introducing a small amount of benzoyl groups (degree of substitution 0.054) or valeryl groups (degree of substitution 0.12) in the lower phase, the partition coefficient of this enzyme could be decreased by more than 100 times. A similar, but weaker, effect on partitioning was observed for bovine serum albumin, lysozyme, lipase and β-lactoglobulin. The partitioning of thylakoid membrane vesicles was strongly affected by the hydrophobic groups on dextran. The membrane vesicles were partitioned toward the phase containing the hydrophobic groups.