EFFECT OF WATER AND ENZYME CONCENTRATION ON THERMOLYSIN-CATALYZED SOLID-TO-SOLID PEPTIDE-SYNTHESIS

We have studied a thermolysin-catalyzed solid-to-solid dipeptide synth esis using equimolar amounts of Z-Gln-OH and H-Leu-NH, as model substr ates. The high substrate concentrations make this an effective alterna tive to enzymatic peptide synthesis in organic solvents. Water content was varied in the range of 0 to 600 mt water per mol substrate and en zyme concentration in the range of 0.5 to 10 g/mol of substrates, High yields around 80% conversion and initial rates from 5 to 20 mmol s(-1 ) kg(-1) were achieved. The initial rate increases IO-fold on reducing the wafer content, to reach a pronounced optimum at 40 mt water per m ol substrate. Below this, the rate falls to much lower values in a sys tem with no added water, and to zero in a rigorously dried system. Thi s behavior is discussed in terms of two factors: At higher water conte nts the system is mass transfer limited (as shown by varying enzyme co ntent), and the diffusion distances required vary. Al low water levels , effects reflect the stimulation of the enzymatic activity by water. (C) 1998 John Wiley & Sons, Inc.