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Aryl amide metallofoldamersas selective saccharide sensors

Periodic Reporting for period 1 - RAMSES (Aryl amide metallofoldamersas selective saccharide sensors)

Reporting period: 2016-03-01 to 2018-02-28

Carbohydrate recognition is fundamental in varied biological processes. It is well established that carbohydrate-binding molecules may have a strong impact on the development of medicinal agents for the inhibition of pathological events such as metastasis and viral/bacterial infections that are mediated by carbohydrate binding, of diagnostic agents for the detection of disease-related glycoproducts, of vectors for targeted delivery and of imaging and therapeutic agents.
However, the design of synthetic receptors for monosaccharides still remains an elusive objective, in particular if recognition is to take place in highly competitive polar media.
The main objectives of this project consisted in the rational design and synthesis of helically folded aromatic oligoamide capsules containing metal ions, in order to achieve high affinity and selectivity for monosaccharides in polar media and to further develop them into sensors.
The devised strategy consisted in an iterative design process that exploits the modular structure of folded synthetic oligomer sequences in conjunction with structural characterization to inform subsequent refinements. The methodology involved achieving the following objectives: a) synthesize a 1st-generation sequence containing the metal-binding monomer and a cavity large enough to encapsulate a guest; b) assess the metal coordination ability of the capsule; c) assess substrate binding ability for a series of relevant guests; d) proposal of a 2nd-generation capsule; e) iterative repetition of c) and d); f) endow the receptor with fluorescent properties.
A 1st-generation oligomer (1) was designed to have a reduced diameter at each extremity and a wider diameter at the centre that includes a metal-coordinating site and was prepared following a well-established procedures.
The results obtained in solution by circular dichroism (CD) and NMR spectroscopies and in the solid state by X-ray crystal structure analysis showed that 1 can coordinate several metal ions leaving part of their coordination sphere available to bind a guest.
Interestingly Mg2+, Ca2+ and Ba2+ binding occurs through 2nd-coordination sphere interactions, i.e. through hydrogen bonds between the capsule inner wall and hydration sphere of the metals. This result expanded the scope of molecular recognition by aromatic helical foldamers.
The ability of the metal-containing sequence 1 to bind carbohydrate guests was assessed in solution using CD and NMR spectroscopies. The results showed that sequence 1 does not bind D-threitol, a 4-carbon carbohydrate, unless it is in the form of a metal complex. In addition, it was found that affinity towards this guest depends on the nature of the metal ion.
The crystal structure of 1-K+/threitol complex validated the complete encapsulation and the involvement of the metal ion in the direct coordination of the guest. The same was found in the case of the crystal structure of 1-Cu2+/threitol complex although here the guest is bound through 2nd-sphere coordination.
To be able to bind larger carbohydrate guests, a 2nd-generation foldamer (2) featuring a larger cavity size was designed and synthesized.
CD titrations showed that 2-Cu2+ can bind various monosaccharide guests in solution and revealed a preference for D-mannose over D-glucose, D-galactose and D-fructose.
An X-ray crystal structure showed that Cu2+ does not bind directly to the guest, but instead solvent molecules bridge the metal ion and the guest. The cavity was found to be too large to efficiently bind D-mannose, which occupies only 43% of the total cavity volume, which agrees with the moderate affinities found for the monosaccharide guests.
A fluorescent monomer was prepared and incorporated in a capsule sequence. Fluorescence spectroscopy titrations showed that an increase in guest concentration causes a decrease in fluorescence intensity, providing proof of concept that a fluorophore can be incorporated on to foldamer capsule sequences and serve as reporter of the binding event.
The results obtained gave rise to two publications in an international peer reviewed journal: Chem. Commun., 2017, 53, 9300 (Featured on inside front cover) and Chem. Commun., 2018, 54, 5078.
Moreover, the work has been presented on several occasions in international meetings: Symposium on Foldamers, Pessac, France, 2016; Conference on Functional Nanocontainers, Tarragona, Spain, 2016 and 12th International Symposium on Macrocyclic and Supramolecular Chemistry, Cambridge, UK, 2017.
The work developed during the project duration went beyond the state of the art of the foldamer field by expanding the scope of molecular recognition by aromatic helical foldamers: a) the discovery that an aromatic oligoamide capsule tightly binds to alkali and alkaline earth metal hydrates through 2nd-coordination sphere interactions, not only was unprecedent in the foldamer field but also revealed a behaviour that sharply contrasts with the classical binding of these metal ions by crown-ether, cryptands or cryptates that act as 1st-coordination sphere ligands. In addition, it was found that magnesium hydrate is bound with very high selectivity in particular with respect to calcium from which it is usually very difficult to discriminate; b) although metals have been used in the foldamer field, for example to build helicates or as modulators of foldamer conformation, the introduction of metal ions into helically folded capsules for the purpose of enhancing binding energies was achieved for the first time. Several cases of selective carbohydrate binding were found, mediated either by direct metal-carbohydrate coordination, or by second sphere interactions between metal-bound water molecules and the carbohydrate guest. The structures of three of these complexes could be elucidated by X-ray crystallographic analysis. These are the first examples of crystal structures of host-guest complexes between carbohydrates and metal-containing synthetic receptors and constitute important milestones in the challenging area of carbohydrate recognition by synthetic receptors, a field that had until recently suffered from a complete lack of solid state evidence; c) the studies performed successfuly provided proof of concept that a fluorophore can be incorporated on to foldamer capsule sequences and serve as reporter of the binding event.
Overall, the project gave a significant contribution to the challenging field of carbohydrate recognition by synthetic receptors. Advancements in this field are poised to have a tremendous impact in the development of medicinal agents for the inhibition of pathological events such as metastasis and viral/bacterial infections that are mediated by carbohydrate binding, of diagnostic agents for the detection of disease-related glycoproducts, of vectors for targeted delivery of imaging and therapeutic agents and of research tools for glycomics. All these potential applications are well in line with the Horizon 2020 societal challenge of “Health, demographic change and wellbeing” and will contribute to the strengthening the Union’s excellence and attractiveness in research and innovation and its economic and industrial competitiveness.
It can also be envisioned that introducing metal ions into helically folded aromatic oligoamide capsules may have been the first step towards other potential applications such as supramolecular catalysis and enzyme mimicry.
Fig. 1 a) metal-assisted guest encapsulation; b) metallofoldamer sensor.