IMMOBILIZATION OF ALANINE AMINOTRANSFERASE BY COVALENT BINDING AND ENTRAPMENT

Alanine aminotransferase was immobilized by covalent binding on two di fferent pre-activated hydroxylic supports and by entrapment in calcium alginate beads. The covalently immobilized derivatives were between 5 and 7.5 times more thermostable than the native enzyme, The apparent denaturation transition temperatures of these derivatives were around 60 degrees C, while those of the native enzyme and the entrapped deriv ative were 44 and 50 degrees C respectively, The covalently immobilize d derivatives resisted unfolding by urea and organic solvents more tha n did the native and entrapped enzymes; they also refolded more effect ively. The pH optimum (7.5) was identical for immobilized and native e nzyme. K-m values of the immobilized enzymes for 2-oxoglutarate substr ate were higher than that of the native, while their V-max values were lower. Immobilized derivatives displayed high catalytic rates and goo d operational stabilities in batchwise synthesis of L-alanine.