Immobilization of Urease by Thiol‐Disulphide Interchange with Concomitant Purification

Urease has been immobilized by covalent attachment to thiolated Sepharose via a mixed disulphide derivative of Sepharose, obtained by reacting thiolated Sepharose with 2,2′‐dipyridyl‐disulphide (2‐Py‐S‐S‐2‐Py). The immobilized urease was used for continuous hydrolysis of urea. The immobilization procedure described accomplished a 6–10‐fold purification of the enzyme preparation used. Since the coupling is reversible, the technique can be used for purification of urease. The chemical selectivity of the mixed disulphide derivative of Sepharose toward urease is discussed with the assumption of close analogy between the mixed disulphide of Sepharose and the reagent 2‐Py‐S‐S‐2‐Py. The reactivity of 2‐Py‐S‐S‐2‐Py, in contrast to other commonly used thiol reagents, appears to be directed preferentially toward thiol groups which are of importance for the urease activity.