Constitutional self-selection from dynamic combinatorial libraries in aqueous solution through supramolecular interactions

Article abstract of DOI:10.1039/c4cc00245h

We describe the predominant formation of a specific constitution arising from the combination of building blocks with different topologies through disulphide chemistry in a Dynamic Combinatorial Library (DCL). The supramolecular interactions established by a zwitterionic cysteine moiety are responsible for the self-selection of one product from all the virtual members of a large library. the Partner Organisations 2014.

Full text of DOI:10.1039/c4cc00245h

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Constitutional self-selection from dynamic
combinatorial libraries in aqueous solution
through supramolecular interactions†
Cite this: Chem. Commun., 2014,
50, 4564
Received 11th January 2014,
Accepted 27th February 2014
Jordi Sola,* Maria Lafuente, Joan Atcher and Ignacio Alfonso*
`
DOI: 10.1039/c4cc00245h
www.rsc.org/chemcomm
We describe the predominant formation of a specific constitution combination of tripodal and bipodal building blocks.8 The stoichio-
arising from the combination of building blocks with different metry and the presence of a template were critical for the latter. More
topologies through disulphide chemistry in a Dynamic Combinatorial recently Sanders has described the diverse topologies obtained by a
Library (DCL). The supramolecular interactions established by a combination of tri- and monothiols in water.9 In order to expand the
zwitterionic cysteine moiety are responsible for the self-selection structural and topological diversity of supramolecular assemblies by
of one product from all the virtual members of a large library.
DCC we decided to study the effect of combining mono-, di- and
trisulphides on the same DCL. In principle, by combining a series of
Self-assembling and self-organization are inherent processes in building blocks of different topology, a broad structural variety is
Nature that occur by the spontaneous folding and assembling of expected. We chose to combine pseudopeptidic C2-symmetric
relatively simple chemical entities.1 For that, the delicate inter- and dithiols 1a–b described earlier by our group10 with the tripodal
intramolecular interactions are established and regulated by proof- building block 28 and a series of monothiols 3a–g. Thus, one may
reading and self-correction.2 Accordingly, the chemical information expect the formation of several compounds with different archi-
stored in a given chemical structure is decoded to render beautifully tectures some of which are schematically represented in Scheme 1b.
complex and fully functional biomolecular systems. For decades,
We started by mixing trithiol 2 with neutral dithiols 1a or 1b at
supramolecular chemists have been fascinated and inspired by this pH 6.5 in an aqueous solution containing 25% DMSO. The use of
biomolecular machinery. More recently, Constitutional Dynamic DMSO allows faster oxidation and exchange within a few hours
Chemistry (CDC) and Dynamic Combinatorial Chemistry (DCC) even at a slightly acidic pH.11 The mixtures were analysed after 24 h
have proposed the implementation of complex dynamic systems and 48 h with no significant changes. Several compounds can be
formed by members able to exchange under thermodynamic detected in accordance with the results published in the litera-
control.3 These dynamic systems (Dynamic Combinatorial ture.8–10 Our DCLs, however, presented less variety of cyclic
Libraries, DCLs) are excellent benchmark models for decoding the products arising from oxidation of dithiols 1a–1b (Fig. 1a, blue
chemical information stored in the combination of simple building trace) as they prefer to form mixed products with 2 (presumably
blocks towards the emergence of new assembled structures.4
Disulphide chemistry has been exploited widely for the genera-
cage-like compounds of formula 1222 and 1322).
To our surprise, when the same experiment was repeated for 1a
tion of DCLs in aqueous media.5 However, most of the systems and 2 in the presence of 2.5 mM of cysteine (enough to saturate all
described are based on molecules with two reacting sites (bipodal). free thiols) a product containing each of the three building blocks
Thus, although some impressive (and somehow unexpected) geo- (1a23a) was predominantly formed (Fig. 1a, red trace). The other
metries have been described such as catenanes and knots,6 the tri- minor detectable species are a combination of 2 with 3 cysteines and
dimensional space covered is relatively small, and the libraries are a structure of the formula 1a223a. Since Ser-derivative 1b presented
mostly directed towards the generation and interconversion of cyclic similar behaviour (Fig. 1b) we focused our studies on Asn-derivative
oligomers.7 The formation of cage-like organic structures has seldom 1a. The distribution of products was mostly unchanged upon varying
been described by oxidation of tripodal building blocks or by the the concentration of Cys from 0.5 mM (equimolar with other
components) up to 100 mM, showing the stability of the product
formed.12 In addition, we combined building blocks 1a and 2 in
different proportions (1 : 2 and 2 : 1) in the presence of 1 eq. of Cys
Department of Biological Chemistry and Molecular Modelling, IQAC-CSIC,
Jordi Girona 18-26, Barcelona, Spain. E-mail: jordi.sola@iqac.csic.es,
ignacio.alfonso@iqac.csic.es; Fax: +34 932 045 904; Tel: +34 934 006 100
per thiol. In these experiments almost all the limiting reagent was
† Electronic supplementary information (ESI): Experimental details, synthetic
consumed in the same product of formula 1a23a whereas the reagent
in excess formed either adducts with Cys or the corresponding
procedures, additional HPLC traces and NMR spectra are available. See DOI:
10.1039/c4cc00245h
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In order to ascertain what structural elements were crucial for the
selection, other monothiols (3b–e) were tested in the libraries. All of
the thiols performed notably worse than Cys in selecting the
corresponding major product. These experiments showed that both
the ammonium and the carboxylate groups are needed in order to
successfully select a particular structure, with the ammonium group
being more critical, as a hydrogen bond donor in that position
clearly enhances recognition. In fact, the experiments done with
+
thiols 3a–e showed the trend NH3 4 OH 4 NHAc (ESI†) to
effectively select a predominant constitution from the mixture.
We turned then to investigate the structure of the multi-
component molecule to see what possible interactions would
lead to the formation of the major product. In principle two
architectures are possible for a molecule of the formula 1a23a:
the symmetrical structure Ia and the less symmetrical Ib
(Scheme 1). We considered Ia more likely as it would present
a more compact structure in solution.
To confirm this possible arrangement we reproduced the synthesis
of 1a23a on a preparative scale. The NMR spectra of the purified
product in buffered water at pH 6.5 with 15% DMSO-d6 were acquired
using water signal suppression by excitation sculpting (Fig. 2a). All the
signals were successfully assigned by a combination of 1H-NMR
and 2D-TOCSY experiments. All expected protons are visible in the
1H-NMR spectra apart from the three protons corresponding to
the alpha carbons of the amino acids from bipodal and tripodal
fragments, which fall in the region of water suppression. However,
these protons are visible in the TOCSY experiment. Three doublets
corresponding to the three amide bonds coupled to the chiral centre
of the amino acids are clearly visible in a 2 : 2 : 1 ratio (HA, HB, HE). In
addition, protons HC and H18 from the phenylenediamine fragment,
and H4 and H2 from the trimesic acid appear as singlets. Overall, this
data is consistent only with the formation of the more symmetrical
product Ia.
Scheme 1 (a) Thiol building blocks used in this study. (b) Schematic
representation of some of the possible architectures generated by the
combination of topologically different building blocks.
Some other structural considerations can be made based on the
1H-NMR spectra. HE displays a relatively low chemical shift compared
to HA, which could be indicative of a twisting out of the aromatic ring
plane. This also explains the large chemical shift difference between
H2 and H4, both from the tripodal aromatic ring. On the other hand,
HC and HB appear in the downfield amide region, which is consistent
with some kind of hydrogen bonding phenomena, most likely with
any of the carboxylate anions present in the molecule. To shine some
light on the possible tri-dimensional structure of the compound in
solution we performed a conformational search using MMFFaq force
field. Molecular modelling rendered a folded structure with several
Fig. 1 (a and b) DLCs formed by 1a,b and 2 (0.5 mM each) in aqueous
solution (25% DMSO, pH 6.5) in the presence (red) or absence (blue) of stabilizing interactions as shown in Fig. 2b.
cysteine (2.5 mM). (c) DLC formed by 1a (0.5 mM) and 2 (0.5 mM) in the
The most stable conformer shows a structure where the pendant
Cys folds over the macrocyclic cavity. The ammonium cation
resides in the centre of the macrocycle and is stabilized by the
presence of L-Cys, D-Cys and homo-Cys (1.0 mM each).
homodimer (ESI†). We attribute the selection of the major product three carboxylates that arise from the trimesic acid moiety. The
formed in the mixture to some sort of self-recognition within the carboxylate group of the cysteine residue would as well be stabilized
building blocks during the assembly process. Thus, stabilization by hydrogen bonding interactions with the macrocycle amide
through supramolecular interactions would lead to the formation of protons (HC/HB). Other hydrogen bonding interactions between
a major, more stable compound. Reversibility tests confirmed the the macrocyclic peptidic amides and the carboxylates would further
reaching of the thermodynamic equilibrium: addition of 3a after 24 h stabilise the structure. Interestingly a cis amide bond is observed
to a sample containing 1a and 2 gave essentially the same mixture as for the tripodal amide that is not part of the macrocycle. We can
the one formed by 1a+2+3a from the beginning (ESI†).
also observe different dispositions (in/out) for the carbonyls of the
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structural differences have an impact on the formation of the major
species (see Fig. 1c). This provides further evidence of a folded
structure in solution as homocysteine would have a looser fit.
We show the importance of the synergic action of subtle
recognition events within the components of a DCL. A combination
of building blocks of different topology could result in a plethora of
possible structures. However, cooperative supramolecular inter-
actions between the building blocks result in the self-recognition
and the selective amplification of a singular constitution even when
it is statistically disfavoured. The structural importance of these
interactions is proved by the difference observed in the chromato-
graphic profile of the DCL when delicate changes are introduced.
We expect that our findings will improve the comprehension of the
behaviour of complex dynamic libraries of compounds.
This work was supported by the Spanish Ministry of Economy
and Competitiveness (MINECO, CTQ2012-38543-C03-03) and EU
(FP7-PEOPLE-2012-CIG-321659). Personal financial support for J.S.
´
(MINECO, Ramon y Cajal contract), M.L. (MINECO, FPI fellowship)
and J.A. (CSIC and European Social Fund, JAE-predoc fellowship)
´
are gratefully acknowledged. We thank Dr Yolanda Perez for helpful
assistance with the NMR experiments.
Fig. 2 (a) Partial 1H-NMR spectrum (500 MHz, buffered H20 with 15%
DMSO-d6, 298 K) showing the aromatic and amide region for molecule
1a23a. (b) Proposed structure for the major product 1a23a (Ia) with atom
labelling (left) and structure obtained by molecular modelling (right).
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4566 | Chem. Commun., 2014, 50, 4564--4566
which is invisible at 254 nm.
This journal is ©The Royal Society of Chemistry 2014