Enzymatic peptide synthesis can be carried out efficiently in solid-to-solid reaction mixtures with 10% (w/w) water added to a mixture of substrates. The final reaction mass contains =80% (by weight) of product. This article deals with acid-base effects in such reaction mixtures and the consequences for the enzyme. In the Thermoase-catalyzed synthesis of Z-Asp-Phe-OMe, the reaction rate is strongly dependent on the amount of basic salts added to the system. The rate increases 20 times, as the KHCO3 or K2CO3 added is raised 2.25-fold from an amount equimolar to the Phe-OMe · HCL starting material. With further increases in KHCO3 addition, the initial rate remains at the maximum, but with K2CO3 it drops sharply. Addition of NaHCO3 is less effective, but rates are faster if more water is used. With >1.5 equivalents of basic salt, the final yield of the reaction decreases. Similar effects are observed when thermolysin catalyzes the same reaction, or Z-Gln-Leu-NH2 synthesis. These effects can be rationalized using a model estimating the pH of these systems, taking into account the possible formation of up to ten different solid phases. © 2001 John Wiley & Sons, Inc.
|Number of pages||8|
|Journal||Biotechnology and Bioengineering|
|Publication status||Published - 5 Jan 2001|
- Acid-base effects
- pH model
- Solid-to-solid enzymatic reactions