Published on Web 10/09/2007
A Peptide Dendrimer Enzyme Model with a Single Catalytic
Site at the Core
Sacha Javor, Estelle Delort, Tamis Darbre,* and Jean-Louis Reymond*
Contribution from the Department of Chemistry and Biochemistry, UniVersity of Berne,
Freiestrasse 3, 3012 Berne, Switzerland
Abstract: Catalytic esterase peptide dendrimers with a core active site were discovered by functional
screening of a 65 536-member combinatorial library of third-generation peptide dendrimers using fluorogenic
1
(
-acyloxypyrene-3,6,8-trisulfonates as substrates. In the best catalyst, RMG3, ((AcTyrThr)
DapArgSerGly) DapHisSerNH ), ester hydrolysis is catalyzed by a single catalytic histidine residue at the
dendrimer core. A pair of arginine residues in the first-generation branch assists substrate binding. The
8 4
(DapTrpGly) -
2
2
) 860 M- min at pH 6.9) per catalytic site is comparable
1
-1
catalytic proficiency of dendrimer RMG3 (kcat/K
to that of the multivalent esterase dendrimer A3 ((AcHisSer)
which has fifteen histidines and five catalytic sites (Delort, E. et al. J. Am. Chem. Soc. 2004, 126, 15642-
5643). Remarkably, catalysis in the single site dendrimer RMG3 is enhanced by the outer dendritic branches
M
8
(DapHisSer) (DapHisSer) DapHisSerNH
4
2
2
)
1
consisting of aromatic amino acids. These interactions take place in a relatively compact conformation
similar to a molten globule protein as demonstrated by diffusion NMR. In another dendrimer, HG3
(
(AcIlePro)
residues is unaffected by the outer dendritic layers. Dendrimer HG3 or its core HG1 exhibit comparable
activity to the first-generation dendrimer A1 ((AcHisSer) DapHisSerNH ). The compactness of dendrimer
8 4 2 2
(DapIleThr) (DapHisAla) DapHisLeuNH ) by contrast, catalysis by a core of three histidine
2
2
HG3 in solution is close to that a denatured peptide. These experiments document the first esterase peptide
dendrimer enzyme models with a single catalytic site and suggest a possible relationship between packing
and catalysis in these systems.
7
Introduction
include cofactor engineering, the installation of cofactors or
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catalytic metal complexes into noncatalytic proteins, and the
search for promiscuous enzyme activities by active site or
The catalytic function of enzymes arises through the relative
positioning of functional groups in space to create a catalytic
site. Designing enzymes de noVo is one of the most challenging
tasks in macromolecular chemistry. Most approaches to create
functional analogues of enzymes are based on modifying the
enzymes themselves or other proteins, for example by selection
for transition state analogue binding, by directed evolution,
or by computational protein design. Artificial enzymes may
also be formed by design or functional selection from folded
linear peptides or noncatalytic proteins. Additional strategies
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substrate modifications.
The dendrimer approach to artificial enzymes follows a
different strategy, belonging to the general theme of synthetic
enzyme models, and proposes to organize building blocks into
a globular macromolecule by means of topology rather than by
folding. Most experiments toward catalytic dendrimers ad-
1
10
2
3
4
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dressed the multivalent display of catalytic groups at the end
of the dendritic branches, which was found in several cases
5
6
1
12
to enhance catalysis by cooperative multivalency effects despite
steric crowding, following earlier reports in polymers with
multiple catalytic and binding groups investigated as hydrolase
mimics. In the context of catalysis in aqueous medium, we
recently reported strong enhancement of the catalytic potency
of histidine and N-terminal proline residues by multivalent
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10.1021/ja074115f CCC: $37.00 © 2007 American Chemical Society