An aqueous two-phase system of polyethylene glycol (PEG) and potassium phosphate was developed for extraction of a cutinase from cell debris of a recombinant Escherichia coli strain. Basic studies to identify the primary factors which affect cutinase partition, namely the influence of polymer molecular weight, polymer concentration and pH were carried out using a purified preparation of the cutinase. The enzyme partition coefficient was enhanced with decreasing PEG molecular weight, increasing tie-line length and pH.