| Budget Amount *help |
¥5,100,000 (Direct Cost: ¥5,100,000)
Fiscal Year 1989: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1988: ¥4,400,000 (Direct Cost: ¥4,400,000)
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| Research Abstract |
The synthetic reactions of amino acid esters and peptides by the catalysis of proteases in hydrophilic (water-miscible) organic solvents were investigated. Attention has been focused on kinetic aspects of these reactions and the effects of immobilization of the enzymes on their catalytic activity and substrate- and stereo-specificities.
The results are summarized as follows: Serine proteases, such as alpha-chymotrypsin, subtilisin BPN', and subtilisin Carlsberg, exhibited catalytic activity for ester synthesis, transesterification, and peptide synthesis. in many of hydrophilic organic solvents which have been considered to deactivate enzymes. Especially subtilisin Carlsberg retains high activity in aqueous ethanol in which the enzyme dissolves. The reaction rate and product yield depend strongly on the nature of solvent, especially water concentration; maximum rate and yield were obtained at water concentration below 10%. The reactions exhibited Michaelis- Menten kinetics, and the kindtic parameters suggest that the rate determining steps for ester synthesis and transesterification are acylation and deacylation of the enzyme, respectively. Furthermore, the reduction of hydrophobic interaction between the enzyme and substrate was suggested compared to that in aqueous solutions. The change in substrate specificity, especially the specificity of amine components, was shown for peptide synthesis in organic solvents.
The large increase in catalytic activity of enzymes was realized by the immobilization to specific support materials, such as chitin, chitosan, and PVA-chitosan blend films.
The results may be interpreted not only by concentration of substrates around the enzymes, but also by activity enhancement of the enzymes by adsorption to the supports.
The results indicate the possibility of controlling enzyme activity and specificity by the selection of media and support materials.
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