Dipeptide binding in water by a de novo designed guanidiniocarbonylpyrrole receptor

Article abstract of DOI:10.1021/ja048587v

A new dipeptide receptor 9, which was designed de novo based on theoretical calculations, efficiently binds dipeptides in water with K_ass > 10⁴ M^-1 by a combination of ion pairing and hydrogen bonds as can be shown by UV-t

Full text of DOI:10.1021/ja048587v

Published on Web 07/01/2004
Dipeptide Binding in Water by a de Novo Designed
Guanidiniocarbonylpyrrole Receptor
Carsten Schmuck* and Lars Geiger
Institute of Organic Chemistry, UniVersity of Wu¨rzburg, Am Hubland, 97074 Wu¨rzburg, Germany
Received March 11, 2004; E-mail: schmuck@chemie.uni-wuerzburg.de
Today, there are only a few artificial receptors known that allow
the complexation of a peptidic substrate in water.1 One reason for
this is that the strength of hydrogen bonds (H-bonds), often
successfully used for molecular recognition in organic solvents,
decreases rapidly with increasing polarity of the solvent, making
the design of receptors for aqueous media challenging.2,3 We
currently explore how additional ionic interactions enhance the
binding affinity of hydrogen-bonding motifs.4 In this context, we
present here a new cationic receptor prototype 9 that efficiently
binds dipeptides in water with association constants Kass > 104 M-1
.
The synthesis of 9 is described in Scheme 1. A Friedel-Crafts
acylation of 2-pyrrole methyl carboxylate 1 with meta nitro benzoyl
chloride 2 using ZnCl2 as the catalyst under kinetic control5 provides
the 2,5-disubstitued pyrrole 3 in 26% yield besides 43% of the 2,4-
regioisomer. The nitro group in 3 was reduced with hydrazine and
Raney nickel to give amine 4 in 92% yield, which was then reacted
with imidazole 2-carboxylic acid 56 to provide the corresponding
amide 6 (80% yield). The structures of 4 and 6 were confirmed by
X-ray analysis. Cleavage of the ester (LiOH, 97%) and subsequent
reaction of acid 7 with mono-boc protected guanidine7 using PyBOP
as the coupling reagent (87%) yielded 8. Deprotection with acid
gave the title compound 9.
Figure 1. NMR complexation-induced shift changes of the amide NHs of
10 in the presence of 9 (40% H2O in DMSO-d6).
Scheme 1. Synthesis of Dipeptide Receptor 9
Receptor 9 was designed de novo based on theoretical calcula-
tions (Macromodel 8.0, Amber*, water solvation)8 to bind dipeptides
with a free carboxylate. The guanidiniocarbonyl pyrrole moiety is
expected to form a hydrogen-bonded ion pair with the carboxylate,9
whereas additional H-bonds between the dipeptide backbone and
the receptor further stabilize the complex. The H-bond from the
imidazole NH can be either neutral (monocation) or partly ionic
(dication), depending on the pH of the solution.
First hints that 9 is indeed capable to bind dipeptides came from
ESI MS experiments which show a distinct signal for a 1:1 complex
between 9 and Ac-Ala-Ala-OH (10) (DMSO/MeOH solution). To
probe the complexation properties of 9 in solution we first
performed NMR titration experiments in 40% water in DMSO.10
Upon the addition of 10 (NMe4+-salt) to 9 (1 mM, monopicrate
salt), significant complexation-induced shift changes can be ob-
served for both the receptor and the dipeptide (Figure 1).11 For
example, the amide NH next to the carboxylate in 10 exhibits a
significant downfield shift with increasing equivalents of 9 added,
whereas the N-terminal amide NH shows an upfield shift (Figure
1). Furthermore, the coupling constants for the amide NHs increase
from 5 to 6-8 Hz upon binding, indicating a more pronounced
â-sheet-like conformation. This is in good agreement with the
suggested binding mode depicted above. Corresponding shift
changes are also observed for receptor 9 (e.g. for the imidazolium
CHs). The linearity of the shift changes not only proves the 1:1-
complex stoichiometry but also shows that even in aqueous DMSO,
complex formation is too strong to measure by NMR. The
association constant for the binding of 10 is therefore estimated to
be Kass > 106 M-1 in this solvent mixture.
The complexation properties of 9 were therefore studied by UV
titration in water (with 10% DMSO added for solubility reasons)
with various dipeptides and amino acids as substrates. The binding
9
8898
J. AM. CHEM. SOC. 2004, 126, 8898-8899
10.1021/ja048587v CCC: $27.50 © 2004 American Chemical Society

C O M M U N I C A T I O N S
Figure 3. Calculated complex structure for the binding of 13 (yellow) by
receptor 9 (gray).
any other dipeptide receptor reported thus far. The general structure
of 9 should also allow the development of a second generation of
receptors with specifically built in side-chain interactions to further
increase the substrate selectivity (for example via an N′-alkylation
at the guanidinium moiety)1b in the future.
Figure 2. Job plot (inset) and binding isotherm for the complexation of
Ac-Ala-Ala-OH (10) by receptor 9 in water (dotted line ) expected UV
change due to simple dilution).
Table 1. Binding Constants of 9 with Various Carboxylates
Acknowledgment. This work was supported by the DFG
(SCHM 1501/2-2) and the Fonds der Chemischen Industrie.
a
carboxylate
K
ass
Gly-Gly (11)
Ala-Ala (10)
Val-Ala (12)
Val-Val (13)
Ala (14)
15.900
30.600
43.800
54.300
7.400
Supporting Information Available: Experimental details for the
synthesis of 9; binding data. This material is available free of charge
via the Internet at http://pubs.acs.org.
Gly (15)
5.200
References
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was followed by the decrease in the absorption of the pyrrole moiety
at λ ) 320 nm (Figure 2) upon the addition of aliquots of the
dipeptide to a solution of 9 (0.01586 mM, chloride salt, 0.5 mM
bis-tris-buffer at pH ) 5.5).12 A Job plot confirmed the 1:1 binding
stoichiometry in water. A nonlinear curve-fitting procedure was
used to determine the binding constants (Table 1). The data show
that 9 binds dipeptides very efficiently even in water with
association constants Kass > 104 M-1, making 9 one of the most
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The dipeptides are bound up to 10 times more efficiently than
simple amino acids (Kass ≈ (5-7) × 103 M-1) for which the
association constants are similar to those for other guanidiniocar-
bonyl pyrrole-based carboxylate receptors, therefore representing
simple ion pair formation.1a,4b Hence, the increase in stability for
the dipeptides must be due to the additional binding sites within
the complex (the H-bonds between the backbone amides and
interactions with the imidazol group). Within the series of dipeptides
studied the complex stability increases, depending on the side chains
present in the order Gly < Ala < Val. This might be surprising at
first glance as there are no specific binding sites for side-chain
interactions present in 9. However, the increase in stability in this
order is in good agreement with both the decreasing flexibility of
the peptide and the increasing hydrophobicity of the side chains.
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favor the formation of â-sheets.13 As the interactions within the
complex with 9 are similar to those found in a â-sheet, it is not
surprising that Val-Val is bound better than Ala-Ala or Gly-Gly,
respectively. Furthermore, within the complex the isopropyl side
chains effectively shield the H-bonds between the backbone amides
from the surrounding solvent (Figure 3) thereby increasing their
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(11) Dilution studies showed that under these conditions no self-association
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(12) Anion binding can be regarded as a beginning deprotonation, as the free
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In conclusion, we have shown here that based on a theoretical
prediction a new and very efficient dipeptide receptor 9 was
successfully realized. The binding properties of 9 are superior to
(14) Chitnumsub, P.; Fiori, W. R.; Lashuel, H. A.; Diaz, H.; Kelly, J. W. Bioorg.
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