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Enzyme-specific activation versus leaving group ability.

de Beer RJ, Bögels B, Schaftenaar G, Zarzycka B, Quaedflieg PJ, van Delft FL, Nabuurs SB, Rutjes FP - Chembiochem (2012)

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.

View Article: PubMed Central - PubMed

Affiliation: Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands.

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Reaction structural pathway of Z-Gly-Act with trypsin.
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fig03: Reaction structural pathway of Z-Gly-Act with trypsin.

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.


Enzyme-specific activation versus leaving group ability.

de Beer RJ, Bögels B, Schaftenaar G, Zarzycka B, Quaedflieg PJ, van Delft FL, Nabuurs SB, Rutjes FP - Chembiochem (2012)

Reaction structural pathway of Z-Gly-Act with trypsin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3569868&req=5

fig03: Reaction structural pathway of Z-Gly-Act with trypsin.
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.

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.

View Article: PubMed Central - PubMed

Affiliation: Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands.

Show MeSH