Chemical approaches to oriented protein immobilization

A procedure has been developed for converting 1,1′‐carbonyldiimidazole‐modified agarose to activated agarose capable of coupling protein in an oriented fashion. This procedure is based on replacing the imidazole groups with orthonitrophenyl derivatives containing ionic and/or hydrophobic groups. The degree of orientation of immobilized chymotrypsin and trypsin was estimated by comparing the activity of both proteases toward low‐ and high‐molecular‐weight substrates. The differences in these activities were greater in the case of enzymes immobilized on agarose gel containing hydrophobic activating agents such as 4‐hydroxy‐3‐nitrobenzophenone and 2‐nitro‐4‐benzylphenol than in those immobilized on gels containing ionic activating agents.