ENZYMATIC-REACTIONS IN AQUEOUS ORGANIC MEDIA .9. DIFFERENCE IN CATALYTIC ACTIVITIES OF SUBTILISIN CARLSBERG AND SUBTILISIN BPN' AND IMMOBILIZATION ACTIVATION FOR ESTER SYNTHESIS AND TRANSESTERIFICATION IN ETHANOL

Free subtilisin Carlsberg (STC) catalyzes the ester formation reaction from N-acetylated aromatic amino acids in alcohols containing small amounts of water. The reaction rate and ester yield are strongly dependent on the water concentration, and the maximum rate and ester yield were obtained at around 2% water. The stereo- and substrate specificities of STC are retained in these reaction media. STC is highly stable in ethanol and activity for ester synthesis was mtaintained for at least 5 weeks at 4°C. In contrast to STC, free subtilisin BPN′ (STB) is inactive for ester synthesis in alcohols, but is markedly activated by immobilization (complexation) to poly(vinyl alcohol) or polysaccharides, such as chitin or chitosan. STC is also an effective catalyst for transesterification of N-acetyl-l-tyrosine methyl ester to ethyl ester. The immobilization-activation of STB was also realized for transesterification. The change in binding of substrates to STC and the activity of the enzyme for esterification, transesterification, and hydrolysis are discussed on the basis of kinetic measurements. © 1990.