Enzyme-specific activation versus leaving group ability.
Bottom Line: Enzyme-specific activation and the substrate mimetics strategy are effective ways to circumvent the limited substrate recognition often encountered in protease-catalyzed peptide synthesis.A key structural element in both approaches is the guanidinophenyl (OGp) ester, which enables important interactions for affinity and recognition by the enzyme--at least, this is usually the explanation given for its successful application.In this study we show that leaving group ability is of equal or even greater importance.
Affiliation: Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands.Show MeSH
Mentions: An appropriate computational technique to study electronic events within or between molecules is ab initio calculations. As the computational requirements are large for these calculations, four compounds were selected: Z-Gly-OGp, Z-Gly-O3G, Z-Gly-NGp, and Z-Gly-OTfe. Furthermore, the system was drastically simplified, in that the enzyme was represented by only the hydroxyl connected to a carbon of the active site serine. In addition, part of the protecting group of the ester was not taken into consideration. This situation is displayed as the white regions of Figure 3 A. The first step of the reaction comprises the formation of a tetrahedral intermediate, which is stabilized by the oxyanion hole (Figure 3 B). The subsequent collapse of the intermediate liberates the alcohol (or amine in the case of NGp) from the complex (Figure 3 C). This was the endpoint of our calculations, as we were interested in differences in leaving group ability. Obviously, the enzyme still needs to be deacylated by a nucleophile to complete the catalytic cycle, which would also proceed by a tetrahedral intermediate.
Affiliation: Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands.