Purification of Lipases and Phospholipases by Heparin-Sepharose Chromatography

In recent years heparin-Sepharose chromatography has revolutionized the purification of enzymes and macromolecules. This method utilizes the specific and electrostatic interactions of heparin with enzymes, growth factors, receptors and blood coagulation factors (1 –3 ). The general scheme of heparin-Sepharose chromatography involves the covalent attachment of heparin to Sepharose. A crude tissue preparation is then applied to the column; only those enzymes and macromolecules that specifically/electrostatically interact with heparin are retained on the column while the other proteins having no affinity for heparin are washed out. The retained enzymes and macromolecules can be eluted from the column either by the addition of an excess of heparin to the equilibrating buffer or by using substances such as salts or denaturants (2 ). Theoretically, the success of heparin-Sepharose chromatography depends largely upon how closely the conditions used in the experiment mimic the native biological interactions.